particle size & zetapotential analyzer model mn401 operation...

Nanotrac Wave

Particle Size & ZetaPotential Analyzer

Model MN401

Operation and Maintenance Manual

OM0022

Rev B

December 2013

OM0022 Contents

12/13 Nanotrac Wave Particle Size And ZetaPotential Analyzer - Operation and Maintenance ii

Copyright, Notices, and Trademarks

Printed in U.S.A. – © Copyright 2013 by Microtrac Inc.

While this information is presented in good faith and believed to be accurate, Microtrac disclaims the implied warranties of merchantability and fitness for a particular purpose and makes no express warranties except as may be stated in its written agreement with and for its customer.

In no event is Microtrac liable to anyone for any indirect, special or consequential damages. The information and specifications in this document are subject to change without notice.

Microtrac® is a registered trademark of Microtrac Inc.

CE Conformity: This product is in conformance with the requirements of the following European Council Directives: 2004/108/EC, the EMC directive, and 2006/95/EC, the Low Voltage Directive. Conformance with any other “CE Mark” directives shall not be assumed.

CAUTION

This is a class A product. In some installations, this product may cause radio interference in which case the user may be required to take

appropriate measures, such as re-locating this equipment.

SYMBOL DEFINITIONS

! This CAUTION symbol on the equipment refers the user to this Manual for additional

information. This symbol appears next to required information in the manual.

WARNING, risk of electric shock. This symbol warns the user of a potential shock hazard where voltages greater than 30 Vrms, 42.2 Vpeak, or 60 Vdc may be accessible.

Protective earth terminal. May be provided for connection of the protective earth (green or green/yellow) supply system conductor.

WARNING, hot surface is present. Refer to Product Manual before removing or opening any covers.

WARNING, laser radiation is present. Refer to Product Manual before removing or opening any covers.

Microtrac Inc. 148 Keystone Drive

Montgomeryville, PA 18936

Sales and Service: (888) 643-5880

OM0022 Contents

12/13 Nanotrac Wave Particle Size And ZetaPotential Analyzer - Operation and Maintenance iii

About This Document

Abstract This manual describes the operation and maintenance of the Microtrac® Nanotrac Wave Particle Size

And ZetaPotential Analyzer.

Revision Notes The following list provides notes concerning all revisions of this document.

Rev. ID Date Notes

A 03/12 Initial release.

B 12/13 Released per ECN ME0519.

References

Microtrac Documents

The following list identifies all Microtrac documents that may be sources of reference for the material

discussed in this publication.

Document Title ID #

Microtrac FLEX Software Operation Manual SW0005

Microtrac FLEX Software Security Features Manual SW0006

Zetrator AutoTitration Accessory - Operation Manual OM0014

Contacts The following list identifies important contacts within Microtrac.

Organization Telephone Address

Microtrac Technical Support (727) 507-9770 Voice

(888) 643-5880

12501-A 62nd Street North

Largo, FL 33773

OM0022 Contents

12/13 Nanotrac Wave Particle Size And ZetaPotential Analyzer - Operation and Maintenance iv

Contents

1. INTRODUCTION ................................................................................................... 1

1.1 Purpose of the Manual .............................................................................................................. 1

1.2 Product Description .................................................................................................................... 1

1.3 Wave Configurations ................................................................................................................. 4

1.4 Safety Information, Cautions, and Warnings ......................................................................... 5

1.5 Specifications .............................................................................................................................. 7

2. INSTALLATION AND SETUP ............................................................................... 9

2.1 Unpacking and Selecting A Location For The Nanotrac Wave ........................................... 9

2.2 Setup for Operation .................................................................................................................. 12

2.3 Connecting And Disconnecting The Nanotrac Wave ......................................................... 12

2.4 Additional Setup Instructions: 'External Probe' Models ..................................................... 13

2.5 Additional Setup Instructions: 'Zetrator' Titration Accessory ............................................ 16

2.6 Additional Setup Instructions: NanoFlex Analyzer ............................................................. 18

2.7 Additional Setup Instructions: MicroVolume Insert for Nano and Zeta Sample-Cells 20

2.8 Final Setup ................................................................................................................................ 24

3. OPERATION ....................................................................................................... 25

3.1 Introduction ................................................................................................................................ 25

3.2 Nanotrac Wave Optical Probes and Sample-Cells ............................................................. 25

3.3 Operation of the Cap, for Wave Models With Internal Sample-Cell ................................. 28

3.4 Operation of the Sample-Cell, for Wave Cuvette Models .................................................. 30

3.5 Using Microtrac 'Flex' Software With Nanotrac Wave ........................................................ 34 3.5.1 Starting The FLEX Software For Use With Nanotrac Wave .................................. 34 3.5.2 Measurement SOPs ..................................................................................................... 36 3.5.3 Measurement SOP Items That Are Specific to Cuvette Model ............................. 37 3.5.4 Measurement SOP Items That Are Specific to ZetaPotential Model ................... 39 3.5.5 Molecular Weight Operation and SOPs .................................................................... 46 3.5.6 Temperature Operation and SOPs ............................................................................ 57 3.5.7 Titration Operation and SOPs .................................................................................... 65 3.5.8 Combining SOPs .......................................................................................................... 67

3.6 Nanotrac Wave Testing Guidelines & Information .............................................................. 68

3.7 Sample Preparation ................................................................................................................. 71

4. PRINCIPLES OF OPERATION ........................................................................... 74

4.1 Introduction ................................................................................................................................ 74

4.2 Physical Principles ................................................................................................................... 74

4.3 Zetapotential Measurement .................................................................................................... 80

4.4 Molecular Weight Measurement ............................................................................................ 81

OM0022 Contents

12/13 Nanotrac Wave Particle Size And ZetaPotential Analyzer - Operation and Maintenance v

5. MAINTENANCE .................................................................................................. 84

5.1 Introduction ................................................................................................................................ 84

5.2 Safe Maintenance Practices ................................................................................................... 84

5.3 Routine Maintenance ............................................................................................................... 84 5.3.1 Cleaning of Surfaces.................................................................................................... 84 5.3.2 Cleaning of Nano or Zeta Sample-Cells and Optical Probes ............................... 84

Without Removing the Cell 5.3.3 Cleaning of Sample-Cells and Optical Probes By Removing the Cell ................. 85

5.4 Troubleshooting ........................................................................................................................ 94

5.5 Requesting Service ................................................................................................................ 101

5.6 Requesting Parts and Accessories ...................................................................................... 101

Tables Table 1-1 Run Time .................................................................................................................. 72

Figures Figure 1: Nanotrac Wave Nano/Zeta Analyzer - Front View ....................................................... 2 Figure 2: Nanotrac Wave Cuvette Analyzer - Front View ........................................................... 2 Significant Features: ................................................................................................................... 3 Figure 3: Positioning the Wave; Remove any packing tape ........................................................ 9 Figure 4: Hardware, Software, and Tools ................................................................................. 10 Figure 5: Accessories and Kits ................................................................................................. 10 Figure 6: Cuvette Types, for use with Wave Cuvette Analyzer Mode ....................................... 11 Figure 7: Rear Panel - Power and USB Connections ............................................................... 12 Figure 8: Unpacking the External Probe ................................................................................... 13 Figure 9: Rear Panel of External Probe Model ......................................................................... 13 Figure 10: Step1: Gaining Access to Set Up the Probe Clamp ................................................ 14 Figure 11: Step2: Remove the Cell Support Cover and Close the Access Cover .................... 14 Figure 12: Step 3: Install the Probe Clamp Shaft into the Cell Support .................................... 14 Figure 13: Step 4: Attach the Clamp to the Shaft, and Attach the Probe to the Clamp ............ 15 Figure 14: Using the Probe Clamp to Raise and Lower the External Probe .............................. 15 Figure 15: Typical Setup for Use With Zetrator ......................................................................... 16 Figure 16: CloseUp View #1 of Tubing Connections ................................................................ 16 Figure 17: CloseUp View #2 of Tubing Connections ................................................................ 17 Figure 18: CloseUp View #3 of Tubing Connections: Flow Cap Is Closed onto the Cell .......... 17 Figure 19: NanoFlex Rear-Panel, showing Probe, External Connections, and Power-Switch. .. 18 Figure 20: Bracket for Raising/Lowering a User's Sample Vessel, ......................................... 19

with Clamp for the Probe and Clamp for the Bracket Figure 21: Opening the Access Cover ...................................................................................... 20 Figure 22: Access Cover Is Open............................................................................................. 20 Figure 23: Gaining Access to Probes and Electrodes (Zeta Model) .......................................... 21 Figure 24: Use Wrench Tool .................................................................................................... 22 Figure 25: Use Probe Holder to Pull Probe and Electrode From Left-Side of the Cell ............... 22 Figure 26: Push MicroVolume Insert In, from Left-Side of Cell, ................................................ 23 Figure 27: Accessing the MicroVolume Insert for Testing ......................................................... 24 Figure 28: Cut-Away View of Nano Sample-Cell ...................................................................... 25 Figure 29: Cut-Away View of Zeta Sample-Cell ........................................................................ 26 Figure 30: External Probe fluid height ...................................................................................... 26

OM0022 Contents

12/13 Nanotrac Wave Particle Size And ZetaPotential Analyzer - Operation and Maintenance vi

Figure 31: Using the Cap and Cap Lifter .................................................................................. 28 Figure 32: Removing the Cap from the Lifter ............................................................................ 28 Figure 33: Removing the Lifter ................................................................................................. 29 Figure 34: Sequence for Cuvette Installation and Removal ...................................................... 30 Figure 35: Sequence for Cuvette Installation and Removal ...................................................... 31 Figure 36: Orientation of Cuvette Surfaces When Inserting Into Sample Cell ........................... 32 Figure 37: Velocity Distribution, Suspended Particles in Brownian Motion ............................... 75 Figure 38: Velocity Distribution, Temperature and Velocity Effects ........................................... 76 Figure 39: Heterodyne Detection of Scattered Light ................................................................. 77 Figure 40: Scattering Efficiency, Full Range ............................................................................. 79 Figure 41: Scattering Efficiency, Rayleigh Range ..................................................................... 80 Figure 42: Gaining Access to the Sample-Cell Area ................................................................. 86 Figure 43: The Sample-Cell Area Under the Access Cover ...................................................... 86 Figure 44: Moving the Cap Out of the Way .............................................................................. 87 Figure 45: Remove the Cell Support Cover (Blue) ................................................................... 87 Figure 46: Description of Cell Support Assembly ..................................................................... 88 Figure 47: Cell Support Components, Nano Model .................................................................. 89 Figure 48: Cell Support Components, Zeta Model .................................................................... 89 Figure 49: Loosen Probe Clamp Screws .................................................................................. 90 Figure 50: Remove Probe(s) From the Cell .............................................................................. 90 Figure 51: Remove Probe(s) From the Cell .............................................................................. 91 Figure 52: Remove Cell for Cleaning ....................................................................................... 91 Figure 53: Reassembly of The Cell, and The 'Gap' Tool .......................................................... 92 Figure 54: Use the Gap Tool to Re-set The Positions of Probe(s) and Electrodes ................... 93

OM0022 Introduction

12/13 Nanotrac Wave Particle Size And ZetaPotential Analyzer - Operation and Maintenance 1

1. Introduction

1.1 Purpose of the Manual

This manual describes the setup and operation of Microtrac's Nanotrac Wave Particle

Size And ZetaPotential Analyzer. This manual also includes information on normal

maintenance, and information on troubleshooting and servicing of the Analyzer.

This manual provides cautions and warnings associated with normal use of the Analyzer.

All users of the Analyzer should be familiar with all cautions and warnings.

This manual is meant to be used in conjunction with SW0005, Microtrac FLEX Software

Operations Manual. There are frequent references to SW0005 throughout this manual.

1.2 Product Description

The Nanotrac Wave is a precise instrument that uses optical techniques, proprietary to

Microtrac, to perform Particle Size ('Size') analysis, ZetaPotential ('ZP') analysis,

Molecular Weight ('MW') analysis of particle samples. The Nanotrac Wave is an easy-

to-use, compact, benchtop unit. To complete the analysis task, the Wave requires a

typical computer with Windows operating system and USB port, and with Microtrac's

FLEX software also installed. The Nanotrac Wave is a low-power instrument,

consuming less than 75W of power under worst-case circumstances, with maximum

options installed.

The industrial design of the Nanotrac Wave makes it compatible for laboratory research,

production quality control, process monitoring / control, and other applications. Nanotrac

Wave analysis can be performed on materials with particle size ranging from nanometers

to microns.

Nanotrac Wave is the next generation of Microtrac particle analyzers that operate on the

Dynamic Light Scattering (DLS) principle. The Nanotrac Wave analyzer is closely

related to Microtrac's Nanotrac and ZetaTrac analyzers, which perform particle size

analysis in the range of 0.8nanometer to 6.5microns, and, for ZetaTrac, also perform

zetapotential measurement.

Figures and descriptions on following pages give additional initial explanation of the

Wave Analyzer. Note that some figures may be representative of the Wave analyzer and

there may some small differences between figures shown and the analyzer that is

delivered.

OM0022 Introduction


Figure 1: Nanotrac Wave Nano/Zeta Analyzer - Front View

(Note: This image may be slightly different than the delivered Analyzer)

Figure 2: Nanotrac Wave Cuvette Analyzer - Front View

(Note: This image may be slightly different than the delivered Analyzer)

Access Cover

Sample-Cell, where

Cuvette is inserted Cap for Sample-Cell

(hinged)

LCD Display

OM0022 Introduction


Significant Features:

* Optical Probe technology. Microtrac's Nanotrac Wave uses well-established probe

technology, proprietary and unique to Microtrac, for determining Size, ZP, and MW

properties of particle samples. Nano models use one optical probe, while Zeta models

use two probes. Solid-state laser-diodes are utilized as light sources for Microtrac's

unique Optical Probe technology. Lasers are typically in 780nm (near-infrared)

wavelength.

* Solvent-resistant, easy-to-clean Teflon Sample-Cells on Internal Probe models. Sample

material to be analyzed is introduced into the sample-cell, where optical probes interact

with the sample to perform the analysis. An optional-use Cell Cap helps protect sample

from contamination during testing, helps keep the Cell clean when analyzer is not in

use, and helps reduce sample evaporation during long-duration tests or if temperature-

control option is used.

* Chemical-resistant outer finish provides easy cleanup from daily use and from most

chemical and sample spills.

* Simple connections and operation - one connection to external DC power, and one USB

cable to connect to the computer (both are provided with Nanotrac Wave). An on/off

switch on rear panel allows laser light-sources to be manually enabled or disabled for

safety, maintenance, or diagnostic purposes.

* After the user manually introduces the sample into the sample-cell, the analysis is

completely automated through use of Microtrac's FLEX Windows-based software. Full

control and adjustment of analysis parameters, as well as selection of multiple data

output formats, are available through the FLEX user interface.

* LCD display provides local indication of basic parameters - sample temperature, total

signal strength, hazard symbols, etc. Data that is presented on the LCD may change

depending on the operation being performed: for example, when 'Background' or

'Loading' operations are selected in the Flex software, the LCD will shift to show a

total-signal-strength indication ( i.e., the 'sample loading' ) coming from the sample in

the Cell. This could be useful when cleaning/flushing the Cell; if Background

operation is selected then the LCD would give a relative indication of how clean the

Cell is ( the lower the signal-strength, the fewer particles there are, therefore the cleaner

the Cell ).

* Principle functions of the Wave are particle size analysis, particle zetapotential

analysis, and particle molecular weight analysis. Options offered with the Wave

include Temperature Control and Flow. Further information about functions and

options are found throughout this manual.

OM0022 Introduction


1.3 Wave Configurations

'ZP' = ZetaPotential 'MW' = MolecularWeight

'Temp.' = TemperatureControl 'Flow' = Flow Cap

Functions Options

Wave Analyzer Configuration: Model, Optical Probe Type, Fiber Type

Sample-Cell Size ZP MW Temp Flow

Wave Nano Configuration,

Optical Probe = Internal

Optical Fiber = SingleMode

Nano

Teflon or

Stainless X



Optical Fiber = MultiMode

Nano

Teflon or

Stainless X X

Wave Zeta Configuration,



Zeta

Teflon

Wave Cuvette Configuration,



Cuvette

X X

NanoFlex Configuration,

Optical Probe = External

Optical Fiber = SingleMode 1

None

X 2 X X



Optical Fiber = SingleMode 1

None

X 2 3 X



Optical Fiber = MultiMode 1

None

X X 3 X

1 External Probe models come in various Probe lengths - contact Microtrac for possible lengths.

2 Molecular Weight function is possible with External SingleMode model, but performance is

downgraded from Internal SingleMode models. 3 Temperature option is possible with Wave Nano External models, but performance is

downgraded from Internal models.

OM0022 Introduction


1.4 Safety Information, Cautions, and Warnings

User Servicing

Any service or repair of Nanotrac Wave analyzer should be coordinated by contacting

authorized Microtrac representative, or contacting Microtrac Technical Support.

WARNING

Do not tamper with or attempt to defeat any safety feature. Use of controls or adjustments, or performance of procedures other than those specified by the manufacturer, may result in hazardous laser-radiation exposure.

Electrical Safety

The Nanotrac Wave‟s external AC-to-DC power supply (provided with the Wave

system), must be plugged into AC power mains with an earth-grounded safety terminal.

WARNING

Use only AC Mains power system with protective-earth ground terminal. Never operate the unit from a power source that does not have a protective-earth terminal. Never attempt to defeat the protective-earth system of the power source.

Note: There are no high voltages present inside the Nanotrac Wave.

Laser Safety

WARNING

Depending on model, the Nanotrac Wave employs either one or two solid-state diode laser sources (IEC 60825-1 Class 3B) of nominal 780 nanometer wavelength with a nominal optical power level of three milliwatts. Laser-radiation is emitted from the apertures of the Optical Probe(s) at <1mW optical power levels. These apertures are accessible to the user of the Wave. All laser-safety precautions should be followed when handling Wave Optical Probes.

Laser safety labels are affixed at appropriate locations on the outside and inside surfaces

of the Nanotrac Wave. Specific laser-radiation apertures are also labeled. Label

examples:

Additionally, whenever the Laser symbol is present on the LCD display, it is an

indication that lasers are on and emitting laser-radiation, and that laser-safety precautions

should be followed at all times.

To avoid all possible exposure to laser radiation, the power switch that is located on the

rear panel of the Wave analyzer can be switched off.

!

!

!

OM0022 Introduction


Temperature Safety

WARNING

Wave analyzers that are equipped with Temperature Control option have surfaces that can reach high temperatures. Under normal operation the user is not exposed to these surfaces, but it is possible for a user to be exposed to surfaces that are hot enough to pose a burn hazard. Users should always be aware of the present temperature of the system.

CAUTION

Wave analyzers that are equipped with Temperature Control option have the capability of heating fluids to as high as +90°C. The user must take all precautions regarding heating of potentially hazardous fluids, including fluid flammability, ignition hazards, vapor hazards. etc.

Whenever the 'Hot Surface' warning symbol is present on the LCD display…

…it is indication that metal surfaces of the Sample Cell Base are at temperatures that can

cause burns to skin, and that precautions should be followed at all times.

If the symbol on the LCD is blinking (this will occur when a surface is hot, AND the

Access Cover is opened), then a burn hazard is imminent and extra precaution should be

followed.

Chemical Safety

Refer to the Specifications section for a list of wetted materials and chemical

compatibility with those materials.

Questions concerning use of Nanotrac Wave with solvents (organic, polar, non-polar),

high-temperature applications, high-concentration acids / bases, etc., should be addressed

to Microtrac Technical Support:

Microtrac Technical Support


Largo, FL 33773

(727) 507-9770

When using any volatile, flammable or caustic material, always use proper and adequate

ventilation, and follow all other safe handling laboratory procedures.

!

OM0022 Introduction


1.5 Specifications

Specifications That Are Common to All Models:

Mechanical Dimensions : ~ 13" H x 15 ½" W x 14 ½" D

Weight : ~ 15 lbs (~ 6.8 kg)

Electrical AC-DC Power Supply : * In: 90-260VAC, 47-63 Hz

* Out: +15VDC±2.5%; 75W max.

Nanotrac Wave : * In: +15VDC In:

With Temperature Control Option: 75W max.

Without Temperature Control: 10W max.

Solid-State Diode Lasers : * Wavelength: 780nm nominal

* Optical Power: 3mW nominal, 5mW max.

* # of Lasers: Without Zeta: 1

With Zeta: 2

Cuvette: 1

Computer Requirements : Refer to Microtrac FLEX Operating Manual SW0005 for minimum system requirements. No special hardware is

required to be added to the computer.

Environmental Ambient Temperature : 10 to 40°C

Humidity : Up to 90%, non-condensing

Sample Conditions for Size Analysis Particle Size Range : 0.8nanometer to 6.5micron

Concentration Range : 0.01% min. to 40% max. (sample-dependent)

Temperature Range1 : +5 to +90°C

Temp. Measurement Accuracy: ±0.1°C

Recommended pH Range : 2 to 12 pH

Sample Conditions for Molecular Weight Analysis Molecular Weight Range : <300Daltons to >20x10

6Daltons

Specifications That Are Specific to Internal Nano or Zeta Models:

Sample Conditions for ZetaPotential Analysis (For Wave With Zeta) Particle Size Range : 10nanometer to 20micron

Electrophoretic Mobility2 : -15.5 to +15.5 μ/sec per volt/cm

Range

ZetaPotential Range2 : -200mv to +200mV

Recommended Conductivity : 0 to 5 milliSiemen/cm

Range Applied Electric Field : Software-adjustable from 1 to 5 kV/m

OM0022 Introduction


Specifications Specific to Internal Nano or Zeta Models (continued):

Sample Volumes, Wave Analyzer, Internal Nano And Zeta Sample-Cells:

Cell Type Material Minimum Volume Maximum Volume3

Nano Teflon ~ 250ul 4 ~ 500ul / 3 ml

Nano Stainless Steel ~ 250ul 4 ~ 500ul / 3 ml

Zeta Teflon ~ 700ul ~ 1.25ml / 3 ml

Temperature Control Option, Wave Analyzer, Internal Nano And Zeta Sample-Cells:

Temperature Control Range : +5 to +90°C

Temperature Control Accuracy : ±0.3°C

Chemical Compatibility, Wave Analyzer, Internal Nano And Zeta Sample-Cells:

Wetted surfaces may include: 316 stainless-steel, oxide films, gold, titanium, chromium,

Teflon, sapphire, Hastelloy C. Recommended suspending medium is aqueous (water); other

fluids (solvents, etc.) can be used; contact Microtrac Technical Support for use with other

mediums.

Specifications That Are Specific to Cuvette Models:

Sample Volumes, Wave Cuvette Analyzer

Cuvette Type Material Minimum Volume Maximum Volume5

Micro Near Polystyrene 50ul Approx. 1ml

Semi Micro Polystyrene 300ul Approx. 2ml

Macro Polystyrene 1,000ul Approx. 3ml

Glass Glass 1,000ul Approx. 3ml

Temperature Control Option, Wave Cuvette Analyzer:

Temperature Control Range : Polystyrene Cuvettes: +5 to +70°C

Glass Cuvette: +5 to +90°C

Temperature Control Accuracy : ±0.3°C

Chemical Compatibility, Wave Cuvette Analyzer:

Polystyrene Cuvettes: Use for aqueous samples. May be suitable for some samples with

nonaggressive solvents. Sample pH range is limited for polystyrene cuvette.

Glass Cuvette: Suitable for all samples.

1 This specification refers to a sample that may be heated or cooled externally to the Wave analyzer. This specification

is primarily intended for models with External Optical Probe, or NanoFlex models with External Probe. That Probe

can accommodate samples with this temperature range. Introducing non-room-temperature samples into the

Internal Sample-Cell or into an installed Cuvette is not recommended. Use Temperature Option to heat or cool

samples in the Cell or in a Cuvette. 2 Mobility and Zetapotential specifications are at nominal sample temperature of +25°C.

3 Maximum Volume values are with / without the Cell Cap in place. Excessive sample volume could cause overflow

when a Cell Cap is put onto the Cell. Without the Cap, excessive sample volume (>3ml) could cause the Cell to

overflow. 4 Minimum Volume of Nano Cell is achieved by adjusting the Volume-Reducing Plug in the Cell.

5 Excessive sample volume could cause overflow if a Cap is placed onto the Cuvette. Without the Cap, excessive

sample volume (>3ml) could cause the Cuvette to overflow.

OM0022 Installation And Setup


2. Installation And Setup

2.1 Unpacking and Selecting A Location For The Nanotrac Wave

The Nanotrac Wave must be operated on a flat and level surface. The surrounding

environment should be such that possibility of sample contamination is minimized. The

selected location should be as free from vibration as possible. If excessive vibration is

present, additional steps should be taken to isolate the Nanotrac Wave. Contact

Microtrac Technical Support for guidance on vibration isolation.

For models with Temperature Control Option installed, airflow is exhausted out through

the bottom surface of the analyzer. For best performance of the Temperature Control

option, the lower rim of the Wave analyzer should be free of obstruction to allow this

airflow.

Some small amount of space is also required behind the Wave. A small external power

supply is used with the Wave. Typically this is placed behind the Wave unit. The

connectors also require some additional space.

The sample-cell area that is beneath the Access Cover may have some tape in order to

secure any moving parts. After unpacking the analyzer, lift the Cover, and carefully

remove this tape.

Figure 3: Positioning the Wave; Remove any packing tape



Included with your Nanotrac Wave shipment are the AC-to-DC power supply, a USB

cable, Microtrac Flex Software, various tools and accessories, and one or more kits of

already-prepared samples for diagnostic testing. Examples are shown in the following

figures. Note: tools and accessories that are provided vary by analyzer model.

1: DC Power Supply 3: Flex Software 5: 'Wrench' Tool

2: USB Cable 4: 'Gap' Tool

Figure 4: Hardware, Software, and Tools

6: Maintenance Kit 7: Disposable Pipettes 8: Diagnostic Sample Kit

Figure 5: Accessories and Kits



9: Macro Cuvette 10: Semi-Micro Cuvette 11: Micro Cuvette 12: Glass Cuvette

Figure 6: Cuvette Types, for use with Wave Cuvette Analyzer Model (These are only provided with Wave Cuvette Analyzer, not with other Wave Models)

9

10

11

9

10 11 12



2.2 Setup for Operation

CAUTION

Before connecting or disconnecting power-supply cable from the Nanotrac Wave, always insure that the power switch, located on the Rear Panel of the Wave analyzer, is turned off. This will minimize possibility of damage to the Nanotrac Wave.

There are several actions that must be taken prior to connection and operation of the

Nanotrac Wave. These actions follow this sequence:

* Install Wave software drivers onto computer. Follow Microtrac Service Instruction

SI-900686-103.

* Install Microtrac FLEX operating software into computer. Follow Microtrac Service

Instruction SI-901107-001.

Once these steps are complete, proceed to the next section.

2.3 Connecting And Disconnecting The Nanotrac Wave

Connect DC power supply and USB cable to Wave's rear-panel connectors as shown.

Connect to AC power source and to USB2.0 port on the PC with Flex Software installed.

Figure 7: Rear Panel - Power and USB Connections



2.4 Additional Setup Instructions: 'External Probe' Models

An example of how External Probe models will be shipped is shown. Remove the probe

from it's packing materials. External probes come with small plastic cap; this should be

retained for future use; it may help to protect the probe when the Wave is not in use.

Figure 8: Unpacking the External Probe

Note that, for external probe models, the probe exits from the rear-panel, and some

additional space is needed behind the unit to allow the probe cable to come around to the

front.

Figure 9: Rear Panel of External Probe Model



External Probe models come with accessories that allow the Probe to be clamped into

place for convenient use with the Wave. Following steps are how to set up the Probe

Clamp..

Figure 10: Step1: Gaining Access to Set Up the Probe Clamp

Figure 11: Step2: Remove the Cell Support Cover and Close the Access Cover

Figure 12: Step 3: Install the Probe Clamp Shaft into the Cell Support



Figure 13: Step 4: Attach the Clamp to the Shaft, and Attach the Probe to the Clamp

The Clamp allows convenient way to manually raise and lower the Probe into a sample

vessel, such as the typical 100ml beaker shown below. It should be noted that for

External Probe models, the Probe Clamp is an accessory; it's use is not mandatory; the

user can hold the Probe using other typical laboratory clamps, and the Probe can be

'dipped' into any sample vessel that the Probe will fit into.

Figure 14: Using the Probe Clamp to Raise and Lower the External Probe

CAUTION

Before installing External Probe into any process with high temperature, high pressure, or very aggressive solvents, consult Specifications, or contact Microtrac, for further guidance.



2.5 Additional Setup Instructions: 'Zetrator' Titration Accessory

Sample titration is possible with Wave system, with the use of the Zetrator titration

accessory. Wave analyzer must be 'Nano' or 'Zeta' model, with 'Flow' option. Example

system setup is shown. All necessary accessories, such as tubing, disposable sample-

cups, etc., are provided with the Zetrator. See further in this manual, as well as manual

OM0014, for more details on operating the Zetrator with the Wave.

Figure 15: Typical Setup for Use With Zetrator

Figure 16: CloseUp View #1 of Tubing Connections



Figure 17: CloseUp View #2 of Tubing Connections

Figure 18: CloseUp View #3 of Tubing Connections: Flow Cap Is Closed onto the Cell

'A': Small pipe, Small tubing, 'To Cell' 'B': Large pipe, Large tubing, 'From Cell'

B

A

B

A



2.6 Additional Setup Instructions: NanoFlex Analyzer

Nanoflex analyzer model uses External Probe, similar to Wave External-Probe model

previously described.

In same manner as with Wave External Probe model:

The Probe exits from the rear-panel of the analyzer;

Connections for power and USB, and the On/Off power-switch, are also located on the

rear-panel.

The NanoFlex Probe can be inserted into various sample vessels, and the Probe's flexible

armor allows many possible setups and positions. If it is preferred, NanoFlex comes with

rear-panel mounting bracket and strain-relief for the Probe; the Probe can be inserted

into the bracket and relief as shown in the following figures:

Figure 19: NanoFlex Rear-Panel, showing Probe, External Connections, and Power-Switch.

As described in previous sections, connect NanoFlex to power supply and USB. Also

follow previous instructions for installation of Microtrac software on the computer. Turn

power-switch 'On'; the LED on front-panel should illuminate.



NanoFlex also provides a sample-vessel mounting bracket, which is positioned on the

front-panel. This bracket has a clamp to hold the Probe; the assembly of the Probe to the

clamp is easy and straightforward. The following figures show how the vessel bracket

can be used to raise and lower the vessel, until the sample is in contact with Probe. There

is an easy-to-use thumbscrew, to clamp the bracket in the 'up' position:

Figure 20: Bracket for Raising/Lowering a User's Sample Vessel, with Clamp for the Probe and Clamp for the Bracket



2.7 Additional Setup Instructions: MicroVolume Insert for Nano and Zeta Sample-Cells

An accessory is available that allow microvolume samples to be measured for size. This

is only available for use with Wave models that have Internal Sample Cell (Nano or Zeta

models).

In order to install and use the MicroVolume Insert, follow these steps:

Open the Access Cover:

Figure 21: Opening the Access Cover

Figure 22: Access Cover Is Open



Loosen the Vent Screw on the Cell Cap. Raise the Cap Lifter and swing it out of the

way.

Remove the Cell Cover. This provides further access to the Probes and Electrodes

(Zeta model is shown here):

Figure 23: Gaining Access to Probes and Electrodes (Zeta Model)



On left-side of Sample-Cell, use Wrench tool to loosen two Probe-Clamp screws for

Optical Probe #2. If Nano model, then only the Plug will held on the left-side of the

Cell:

Figure 24: Use Wrench Tool

Use the Probe Holder to pull Probe and Electrode from the left-side of the Cell (Zeta

model).

Figure 25: Use Probe Holder to Pull Probe and Electrode From Left-Side of the Cell

*Loosen 2 screws…

1

Tool

2x X

X X

X

X

X

*…use Probe Holder, and pull a short distance

to remove Probe 2 and

Electrode…

Probe Holder

2

X



Leave the #2 Probe and Electrode clamped in the Probe Holder, and leave it to safely

rest so that nothing can be damaged. Push the MicroVolume Insert into the forward

hole of left-side of the Sample-Cell:

Push the Insert in, across the Cell, until the tip of the Insert is in contact with the face of

Probe #1 (Zeta model). With micropipette (not supplied with Wave analyzer), the

user can carefully insert 15 to 20ul of clean fluid (for SetZero) or sample (for Run),

into the micro-well of the Insert:

Figure 26: Push MicroVolume Insert In, from Left-Side of Cell, Until It is Contacting the Surface of Probe 1.

*Probe and Electrode should not be removed from Probe Holder.

*Push MicroVolume insert through the Sample-Cell.



With MicroVolume Insert in place, do not replace the blue Cell Cover. Swing the Cap

Lifter back over the Cell and push down to put the Cell Cap in place. Close the

Access Cover.

Now, the micro-well of the Insert is accessible, and with micropipette (not supplied),

the user can carefully insert and remove samples.

For best results, the user may wish to clean the Insert between each sample. After

extracting the sample from the Insert, the Access Cover is opened and the Insert is

pulled from the Cell, cleaned, and re-inserted. To make this process easier, the user

may decide to remove the Cap Lifter, as shown in previous sections of this manual.

Then the Lifter will not be in the way. The Access Cover must be down and in place,

to complete it's interlock, before any measurements can be made.

Figure 27: Accessing the MicroVolume Insert for Testing

2.8 Final Setup

It is recommended that after power-switch (located on rear-panel) is turned on, that about

15 minute warm-up period should be allowed, to let all components of Wave analyzer

stabilize, then the Wave should be ready to use. Wave readiness is indicated on the LCD

display.

OM0022 Operation


3. Operation

3.1 Introduction

A complete Nanotrac Wave Particle Size Analyzer system consists of the Wave

Analyzer, a computer with Windows operating system, power-supply, and USB

connecting cables. The Microtrac FLEX Software, installed on the computer, provides all

operator interface functions, data acquisition and analysis commands, and data report

formatting, as well as database data retrieval and supervisory control.

FLEX Operating Manual SW0005 contains detailed information on the use of FLEX to

control the Nanotrac Wave adjust analysis parameters, save and recall data, etc. The

user should refer to this manual for details on Flex and it's features. Excerpts of SW0005

are given in this section of this document to help the user complete initial installation and

to begin using the analyzer.

3.2 Nanotrac Wave Optical Probes and Sample-Cells

'Nano' and 'Zeta' Models

The Nanotrac Wave performs particle size analysis and particle zetapotential analysis by

the principle of Dynamic Light Scattering (DLS). The purpose of the optical probe

assemblies is to deliver laser-light to the sample, and simultaneously to collect the portion

of this light that is scattered back from the sample's particles.

For Nano and Zeta models, the Wave's optical probes are horizontally mounted in the

sample-cell. The sample cell is mounted flush with a user-access-cover, to allow easy

cleaning if spills should occur. Sample is introduced into the sample area of the cell.

The cutaway view demonstrates the minimum sample required to perform an analysis -

the probe surfaces must be completely submerged. This allows for an absolute minimum

of sample to be used per analysis-run. Maximum amount of sample should be limited

such that the cell does not overflow.

Figure 28: Cut-Away View of Nano Sample-Cell

OM0022 Operation


For Nano models, a stainless-steel 'plug' is also installed horizontally in the sample-cell,

opposite from the Probe. This plug's position can be adjusted, to further reduce the

amount of sample that is required.

Figure 29: Cut-Away View of Zeta Sample-Cell

For Zeta models, two electrodes of stainless-steel and teflon are also installed

horizontally in the sample-cell, opposite from each Probe. These are used during the

zetapotential portion of the analysis. Note that the electrodes are paired with the optical

probes, and note the gap between the probes and the electrodes. This configuration is

crucial to proper zetapotential analysis operation.

'External Probe' and 'NanoFlex' Models

External Probe models do not use a Sample-Cell. Instead, as previously mentioned,

External Probes can be 'dipped' into many types of sample vessels that a customer can

choose. For best results, keep the face of the probe a minimum distance from the bottom

of the vessel. And, the Probe may have a 'seam' where it's parts are joined. Do not

immerse the Probe beyond this seam or it may damage the Probe.

Figure 30: External Probe fluid height

~0.25"

Do not immerse

beyond the 'seam'

External Probe

OM0022 Operation


'Cuvette' Models

Previous sections of this manual describe types of cuvettes that can be used with Cuvette

Model of Wave analyzer. In the case of Cuvette model, the cuvette is the sample-cell.

Unlike other Wave analyzer models, the Optical Probe does not enter or touch the

sample.

Minimum volumes that must be used with various cuvette types are found in the

Specifications section of this manual.

All Models

All Wave analyzer configurations have an integrated temperature sensor for monitoring

sample fluid temperature. This allows measurement of sample temperature with an

accuracy of +/- 0.1ºC.

OM0022 Operation


3.3 Operation of the Cap, for Wave Models With Internal Sample-Cell

The built-in Sample-Cell of Nano and Zeta models are provided with a Cap and Cap

Lifter. The Cap has several purposes:

* Keep out contamination during sample test runs;

* Reduce the rate of evaporation when the Temperature Option is used to heat samples;

* Keep the Cell clean when the system is not in use.

Operation of the Cap is simple:

1: Use the Lifter to extract the Cap 2: Swing the Cap out of the way, to have

from the Cell access to the Cell

Figure 31: Using the Cap and Cap Lifter

The Cap can be removed from Lifter, for cleaning or replacement:

Figure 32: Removing the Cap from the Lifter

OM0022 Operation


Use of the Cap and Lifter is optional; both can be removed if preferred. If preferred, the

Cap alone can be used, as shown:

Swing the Lifter clockwise 90°, The Cap can be used by itself

then lift it up and out

Figure 33: Removing the Lifter

The Cap has some features:

* The small screw at the top is a vent, to help prevent possibility of the Cap being air-

locked to the Cell.

-Before the Cap is lowered onto the Cell, loosen the vent slightly by turning

counterclockwise (CCW). This will allow air to escape and prevent pressurizing

the Cell. Once the Cap is in place, close the vent by turning clockwise (CW) until

the vent is hand-tight. Do not over-tighten.

-Before the Cap is lifted from the Cell, loosen the vent slightly by turning CCW. This

will allow air to enter and the Cap to be released from the Cell.

-For most processes, especially operation at room temperature, use of the vent is

probably not necessary, as the Cap should insert and remove from the Cell anyway.

* An o-ring around the perimeter of theCap helps to reduce evaporation during

Temperature testing. This o-ring can be replaced if it becomes worn.

A special version of the Cap provides the Flow option. The Flow Cap is installed onto

the Lifter in the same way as Nano/Zeta Cap. See previous section in this document for

more information, to setup the Flow Cap.

OM0022 Operation


3.4 Operation of the Sample-Cell, for Wave Cuvette Models

The Wave Cuvette analyzer has a hinged cover over the sample-cell, where a Cuvette is

inserted. The cover has several purposes:


* Keep the Cell clean when the system is not in use.

Note the orientation of the different Cuvette types, from the following figures. Operation

of the cover, and inserting / removing Cuvettes is simple:

Figure 34: Sequence for Cuvette Installation and Removal

Cuvette Cell, with Hinged Cover

..to expose the Cuvette.

Lift the Cover..

OM0022 Operation


Figure 35: Sequence for Cuvette Installation and Removal

Cuvette can be inserted or removed

Front Face

Right Face

A light push will 'click' the Cuvette into final position

Note 1: Micro Cuvette (shown here) will not protrude as far as the other types of Cuvettes

Note 2: Close the Cover after the

Cuvette is installed

OM0022 Operation


9: Macro Cuvette 10: SemiMicro Cuvette

11: Micro Cuvette 12: Glass Cuvette (shown with Cap)

Figure 36: Orientation of Cuvette Surfaces When Inserting Into Sample Cell

9

10

11 12

Front Face

Right Face has orient. mark and

clear window

Front Face

Right Face has orient. mark and

clear window

Front Face has orient. mark that

points to the Right Face

Right Face has clear window

Front Face

Right Face has 'G'

mark and clear window

OM0022 Operation


Cuvettes can be used with or without Caps. Cuvette Caps are also provided with the

analyzer. There are two types of Caps that can be used: plastic and silicone-rubber.

The Caps have several purposes:


* Keep out contamination during short-term storage of samples when not being tested in

the Analyzer;

* Reduce the rate of evaporation when the Temperature Control Option is used to heat

samples. It is recommended to use the silicone-rubber Cap when operating the

Temperature Control Option; this will further help keep the sample from evaporating.

OM0022 Operation


3.5 Using Microtrac 'Flex' Software With Nanotrac Wave

FLEX Operating Manual SW0005 contains detailed information on the use of FLEX with

the Wave Analyzer. The user should refer to this manual for further details on Flex and

it's features.

3.5.1 Starting The FLEX Software For Use With Nanotrac Wave

Start the Main FLEX Software Window, from either the Windows Desktop icon

or from the Windows 'Start' button.

In Menu Bar at the top, click 'Tools' - 'Hardware Configuration':

OM0022 Operation


Dialog-box will open; go to 'Nanotrac / Zetatrac/ Wave' tab, under 'Bench Type', select

'Nanotrac Wave'. IF the Wave is also being used with Zetrator titration accessory, then

select 'Installed', and choose the serial COM port that the Zetrator is connected to (further

information about Zetrator setup can be found in manual OM0014):

Click 'OK' to save these selections and to close the dialog-box.

This operation only has to be performed once. If the user does not change the Bench

Type, then every time that Flex is opened, it will choose the Wave as the DLS Bench

Type.

Now from the Menu Bar of the Main Flex Window, click 'Measure' - 'Select Instrument',

and select 'Nanotrac Wave':

OM0022 Operation


If all software and drivers are correctly installed, and then if a valid Wave analyzer is

connected to the PC, then Flex should display the Measure Window:

3.5.2 Measurement SOPs

Measurements are set up from the SOP panel, accessed from the Toolbar:

OM0022 Operation


Further information about Measurement SOPs can be found in Flex Software Manual

SW0005.

3.5.3 Measurement SOP Items That Are Specific to Cuvette Model

If Wave analyzer is Cuvette model, there will be additional choices in the Measurement

SOP.

When 'SOP Options' panel is opened, under 'Analysis' tab, will be 'Cuvette' tab. This

allows the user the ability to select which of the four types of cuvettes to use for this

SOP:

Also in 'SOP Options' panel, under 'Timing' tab, is available selection to 'Enable

AutoZero':

OM0022 Operation


If the 'Enable AutoZero' box is checked, then when the Measurement SOP is closed and

returns to the Flex Measurement window, the 'AutoZero' annunciator will be shown at the

'SetZero Status' area:

If the 'Enable AutoZero' box is not checked, then the SetZero Status will show the date

and time that the last valid SetZero was performed:

If no valid SetZero has been performed, the SetZero Status annunciator will be red,

indicating that either SetZero must be performed, or 'Enable AutoZero' must be checked.

OM0022 Operation


Cuvette model of Wave analyzer is the only model that has AutoZero option. This is

different from other Wave models; other models require a valid SetZero to have been

performed prior to any sample Runs. And, in other models, if Flex software is closed,

that valid SetZero is 'lost'; so when Flex is re-started, the SetZero Status is again red,

meaning that SetZero must be performed. The AutoZero option with the Cuvette

analyzer allows samples to be Run as soon as the Flex software is started; there is no

SetZero to be performed, and there is no SetZero that is 'lost' when Flex software is

closed.

Cuvette models come with a SetZero that has already been performed, and the SetZero

data is stored in permanent memory inside the Cuvette Wave electronics. This is the

AutoZero selection; if the AutoZero selection is checked, the data from that original,

stored, SetZero will always be used.

It is for the user to determine if they wish to use AutoZero or not. With the three types of

plastic cuvettes (Macro, SemiMicro, Micro), it may be better to use AutoZero. These

types of cuvette are disposable. For the Glass cuvette, which will be cleaned and re-used,

it may be desirable to perform SetZero with clear fluid, to account for any changes to it's

background (SetZero) as it is cleaned and used multiple times.

3.5.4 Measurement SOP Items That Are Specific to ZetaPotential Model

Additional information for using Microtrac Flex software to measure Zetapotential can be

found in Flex Software Manual SW0005.

ZetaPotential Measurement - Setup Parameters

For Wave analyzers with Zeta option installed, Zeta Potential measurement is set up in

'SOP' - 'Options' - 'Zeta Potential' tab:

Check the 'Enable Zeta Potential' box to instruct the instrument to include the Zeta

Potential function when sample measurements are performed. If this item is NOT

checked, then only the particle-size measurement function will be performed.

Enter the Dielectric Constant for the suspending fluid. This value is required for the Zeta

Potential calculation. Default is 79, the dielectric constant of Water.

OM0022 Operation


Click 'OK', then 'Close' to close the Measurement Setup Options dialog, and to save the

ZetaPotential settings. Note that there is now an enunciator on the Measurement

window, telling the user that ZetaPotential measurement function is enabled.

When a zeta run is completed, zetapotential data is presented on an individual 'tab' of the

main Measure Window:

OM0022 Operation


When Zeta Potential measurements are “Enabled” the measurement process contains

additional data collections that consume more time than a Particle-Size-only

measurement. Both Particle Size and Zeta Potential are measured and reported when

Zeta Potential Measurements are “Enabled”.

Total measurement time will be:

'Zeta' Disabled (Sizing Only)

Total SetZero Time = SOP 'SetZero' Time

Total Run Time = SOP 'Run' Time

'Zeta' Enabled (Sizing Plus Zeta)

Total SetZero Time = SOP 'SetZero' Time * 2

Total Run Time = (SOP 'Run' Time * 2 ) + (Additional 20 to TBD seconds)

For example: an SOP with SetZero Time = 30-seconds and Run Time = 120 seconds,

with Zeta Potential enabled:

Total SetZero Time = 30 * 2 = 60 seconds

Total Run Time = (120 * 2 ) + (Additional 20 to TBD seconds) = 240 + (TBD) seconds

The additional time required (TBD), for a run with zeta, is dependent on sample; some

samples may take longer to make final determination of zetapotential value.

Therefore, in order to get best possible data in shortest amount of time, the selection of

SOP parameters is as important for zetapotential as it is for sizing.

Performing Zetapotential Measurements

Add clear fluid to the sample cell and click the „BKG‟ toolbar button, to perform a

background check. This check can be used to determine the level of particulate in the

cell prior to the start of a SetZero or a measurement Run. A Background Check screen

appears; a desirable goal is to have Background ≤ 0.01 prior to SetZero or Run.

Zeta Background Check Screen - Additional cleaning is needed

(Note: Shown as an example; actual Flex software and calculated data may differ)

OM0022 Operation


Zeta Background Check Screen - Clean, ready to perform SetZero or Run

(Note: Shown as an example; actual Flex software and calculated data may differ)

The Setzero (Background Measurement) is performed by clicking on the FLEX

toolbar. Note the color of the SetZero enunciator on Measurement window:

* Yellow with „No SetZero‟: Remains like this until a successful SetZero is completed;

* Green with Date and Time: A successful SetZero has been completed;

* Red with Date and Time: SetZero attempt was not successful; run Background Check.

When the setzero is successfully completed, extract the fluid if desired, to prevent sample

dilution. Add sample material to the sample cell; add sufficient sample to immerse the

optical probes.

Click the SOP toolbar button to adjust Measurement Parameters, Sample IDs, etc, if

desired. Refer to Flex Software manual SW0005 for guidance on setting parameters.

ZetaPotential - Introducing Sample to the Analyzer and Loading Check

Click on „LD‟ toolbar button to launch the Nanotrac Wave zeta loading screen. Note that

this is a measure of sample concentration. Accurate refractive indices for particle and

fluid are necessary for correct calculation of concentration here. When the „loading bar‟

is within the acceptable „green zone‟ limits, and the Status enunciator indicates „Ready‟,

it is ready to proceed with a measurement run, which will use the previously-entered

parameters. If sample concentration is too high, the loading bar will out of the green

zone, and the Status should show „Dilute‟, indicating the action to be taken. If

concentration is too low and the loading bar is below the green zone, the Status indicator

should show „Add Sample‟.

OM0022 Operation


ZetaPotential - Performing the Measurement ('Run')

Click to perform the Particle Size and Zeta Potential measurement.

Complete the Optional Zeta Potential Sample Information form (shown below), and click

to perform the particle-sizing and zetapotential measurement functions.

OM0022 Operation


ZetaPotential - Sample-Information Fields

The information in these fields DO NOT directly influence Zeta Potential

measurements and are for informational purposes only.

Fluid ID: Enter an Identifier for the Fluid (carrier) (this is a text field)

Dispersant ID: Enter Identifier for the Dispersant ID (if used) (this is a text field)

Dispersant PH: Enter the sample pH (this is a number field)

Dispersant Concentration: Enter the dispersant concentration (this is a text field)

Particle ID: Enter an Identifier for the Particle (this is a text field)

Sample Concentration: Enter the sample concentration (this is a number field)

NOTE 1: Any previously-entered values will be kept and shown for each future

sample. Be sure to change these entries as required for each sample.

NOTE 2: Clear any unwanted sample information from this form if no information

is desired. The information in this form WILL be saved and displayed

with the results of the measurement. All fields can be cleared

simultaneously by clicking the button.

NOTE 3: Particle Size Data will be displayed PRIOR to the Zeta Potential

measurement being performed.

Click 'Continue' and the measurement run will begin. Throughout the run, the progress

of the measurements is indicated in the Measurement window.

NOTE: Zeta Potential Sample Information can be changed from the completed data

display (measurement or database recall) by clicking the button. This will open the

Zeta Potential Sample Information dialog, and if you exit by clicking the

button, the data display will be updated to the new values. If the data has been saved to a

database record, then it is the user‟s responsibility to “Update” the full record, to

reflect the zetapotential sample information changes that were made.

ZetaPotential - Selecting Data Reports

Reports that include Zeta Potential data are selected from the “Reports Select/Design”

dialog box. See Flex Software Manual SW0005 for information on customizing report

content.

For ZetaPotential data to be presented in printed reports, several pre-designed reports

have been provided. The user must select one or more of these reports for printed

ZetaPotential results to be produced. To select one or more of these reports, click the

'Reports - Select/Design' menu.

Select one of the reports whose name contains 'Zeta', and click 'Add', to add this format to

the 'Selected Reports'.

OM0022 Operation


OM0022 Operation


3.5.5 Molecular Weight Operation and SOPs

Additional information for using Microtrac Flex software to measure Molecular Weight

can be found in Flex Software Manual SW0005.

Molecular Weight - Using Microtrac Flex Software's Control Panel

The Molecular Weight Panel is used to:

* Input user-supplied information used to calculate Molecular Weight.

* Measure a series of samples of different, known-weight concentrations.

* Calculate Molecular Weight based on Debye Plot data collection technique.

* Calculate Molecular Weight based on Hydro-Dynamic Radius data collection

technique.

* Calculate Molecular Weight as an Average of a number of data points. (Only applies

when sample Weight Concentrations do not vary enough for accurate Debye

Plotting.)

To open the Molecular Weight Control Panel Click the MWt button on the Measure

toolbar:

The MW Control Panel is launched:

Note: for convenience some common DLS Analyzer functionality (SOP, ID) can be directly accessed from this control panel.

OM0022 Operation


Steps to follow in the Molecular Weight Control Panel are marked in the panel as Step1

through Step 4. These steps MUST be followed “In Order”. Each Step is disabled until

the previous step is performed.

Step 1: Select the Calculation, and Prepare for measurement:

* Prepare 2 to 5 samples at various molecular concentrations (see Sample Preparation

for Molecular Weight Analysis, later in this section). Highest concentration must be

greater than 1.1 times the lowest concentration before a 2nd

Virial Coefficient will be

calculated. A range from C=.002 to C=.05 is usually sufficient.

* From the Molecular weight control panel, “Select Calculation(s)”, pull down the list,

and select the calculation:

- 'Debye' for just Debye MW;

- 'Hydro' for just Hydrodynamic MW;

- 'Debye and Hydro' for both Debye plot MW and Hydrodynamic MW.

Step 2: Enter Particle And Fluid Parameters:

* Under Step 2, some parameters about the fluid and the particle must be entered.

Parameters depend on the calculation(s) that were selected in Step1:

- If only 'Debye' calculation is needed, then user enters Differential Index of

Refraction, also known as Incremental Refractive Index; designation is 'dn/dc'.

- If only 'Hydro' calculation is needed, user enters the molecular particle's density.

- If both 'Debye and Hydro' calculations are needed, user enters both parameters.

* Notes:

- If the density is not known, it can be measured using the procedure Measuring the

Molecular Density, this is found later in this section.

- If the Incremental Refractive Index is not known, it can be measured using the

procedure Measuring dn/dc With Microtrac Refractometer Software Utility,

this is found later in this section.

OM0022 Operation


* The Molecular Weight Control Panel also allows direct access to some functions in

order to optimize the MW data collection. In SOP, select the optical parameters for

the molecular suspension, and select a run time. Typically 100 seconds run time is

good for most molecular samples. Also, appropriate sample ID1 and ID2 can be

entered. ID1 should describe the molecular species. It appears as the title of the

Debye plot.

Step3: Collect the data:

* Enter the concentration value C of the lowest concentration sample on the first line in

the “Collect Data Points” table. Samples are to be run consecutively in ascending

order of concentrations. C can be entered for all samples at the start of the MW

measurement, or one-at-a-time before each run.

- For further explanation of sample concentration, see Sample Preparation for

Molecular Weight Analysis, found later in this section.

* Load the sample cell with 1-2ml of the first sample. For best MW results, it is

important to first “condition” the cell to the new concentration. Load the cell with an

initial 1-2ml, stir, remove and discard this sample. Add a 2nd

1-2 ml of sample. Any

loss of concentration to the walls and surfaces will have taken place and the cell will

be conditioned for the second loading. Note: If running 'Hydro' only, then load the

sample cell with a low concentration molecular suspension. Because of molecular

interactions, the hydrodynamic size is dependent on concentration. The hydro MW

will be calculated for all samples that are run when “Hydro” is selected. The MW

calculated from the lowest concentration should be selected to represent the

Hydrodynamic MW.

* Click 'Run'. After the run time the size distribution and light intensity of the molecular

mode will be determined and entered in the table. KC/R is calculated and entered in

the table and the point is plotted at the appropriate C on the Debye plot graph.

* Repeat this data-taking procedure for each of the prepared samples. Use the cell-

conditioning step for each sample, for best MW results. Up to 5 points can be

plotted.

OM0022 Operation


Step 4: Calculate the Results:

* After all samples are run, a linear data-fit is implemented for the measured points by

clicking on “Calculate”. The Molecular Weight and 2nd

Virial Coefficient are

calculated and displayed in the top table along with the MW measurement

parameters.

OM0022 Operation


Optional: Print a report by clicking the button.

Molecular Weight - Debye, Without The 2ND Virial Coefficient

Molecular Weight can be measured without the 2nd

Virial Coefficient, by using two or

more samples of the same concentration, or with concentrations less than 1.1x apart.

Such samples will have a MW calculated from the average Rayleigh Ratio of the points.

For accurate MW value the samples should have low concentrations.

Procedure (follows the above procedure for Debye Plot measurement):

* Prepare 1 sample at a low concentration (C < 0.005 g/mL).

* On the Control Panel, enter C into the table, for 1st measurement (Row 1, under 'Wt.

Conc.').

* Initiate the first run.

* Without changing the sample enter the same C into the table for second measurement.

* Initiate the second run. At the end of the run the Debye plot will have two points at the

same C.

* Click on the step 4 “Calculate” button. A message will come up indicating that the 2

concentrations are too close to determine 2nd

Virial. Press “Continue”.

* Molecular Weight will be calculated from the average of the two points and displayed

in the “Calculated Results” table.

Molecular Weight Data Display and Data Retrieval

When a Molecular Weight experiment is completed, the data is also placed in the

Molecular Weight Data Display. A button will appear in the Measure Window menu bar

to open this display. The data will remain in this display until a new Molecular Weight

experiment is performed.

OM0022 Operation


If Molecular Weight Data Collection is performed with 'Database Save' enabled, then the

data can be recalled from the database to the Molecular Weight Data Display. Database

records that are part of an MW experiment are uniquely labeled for easy identification.

When a Molecular Weight experiment is performed, a unique numeric ID is assigned to

the DB record for each Run in the Experiment. This ID number, preceeded with the

prefix “Mole Wt:”, replaces the SOP Title item. See Title column below in the Query

Results tab of the Database Retrieval Display.

To retrieve the data from a saved MW experiment:

1. Select all the records that have the same IDs.

2. Right click on the display and select “Retrieve to Molecular Weight Data Display”

to reproduce the molecular weight data as shown above.

Discarding Invalid Data Points:

If a Molecular Weight experiment produces an invalid data point, the user can choose to

“Discard” that data point at any time and re-calculate the Molecular Weight without that

data point being included.

Typically if the Correlation Coefficient is < 0.9 then one or more data points is most

likely invalid. The Molecular Weight Results data item for Correlation Coefficient will

display with red text and a yellow background for correlation values < 0.9 (see below).

OM0022 Operation


In the example above, data point 3 on the Debye Plot is the invalid data point. This data

point can be “Discarded”:

1. “Un-check” the Check Box in the “Sel” column of the Data Points display for the

desired point to discard.

2. Results are “Automatically Re-Calculated” producing the results shown below. NOTE

that the discarded point on the Debye plot is indicated by an “X” on that data point

and that the Correlation Coefficient is now >= 0.9.

OM0022 Operation


Molecular Weight - Measuring dn/dc With Refractometer Software Utility

In order to determine Molecular Weight from the Debye Plot, the value of the differential

index of refraction, dn/dc, is required for the molecular solution being measured. One

example of published literature in which values of dn/dc are tabulated, as determined

from refractometry, is the volume:

M.B. Huglin, in Light Scattering from Polymer Solutions, M.B. Huglin, Ed., Academic Press, 1972; Ch. 6.

Microtrac‟s Molecular Weight application also provides a means of measuring dn/dc by

utilizing the magnitude of the Fresnel reflectance from the sapphire window of the

Microtrac's DLS Analyzer optical probe. The light reflected at the interface of two

dielectrics is proportional to the square of the ratio of the index-difference to index-sum

of the two dielectrics:

2

333.175.1

333.175.1)/(

Areflected NIntensity

sapphirewater (7)

2

75.1

75.1)/(

solution

solutionAreflected

n

nNIntensity

sapphiresolution (8)

The Analyzer probe focuses an optical laser source at the interface of the sapphire

window (index = 1.75) and the molecular suspension. The reflected light is collected by

the focusing optics and delivered through an optical fiber to a detector. In operation, the

light collected from a known water/sapphire reflectance is compared with the light

collected from a molecular suspension of a known concentration C. From these two

measurements the index of the suspension is determined and the value of dn/dc is

calculated using the input concentration value, Cwv, of the suspension.

Procedure for using dn/dc Utility:

Clean the sample-cell in accordance with procedures in this manual. Flush the sample

cell with DI water until clean, then leave DI water in the cell.

* From MW Control Panel, Step 2 'Parameter Entry', click on “Measure”. This launches

the dn/dc 'worksheet':

OM0022 Operation


* Follow prompts; and in the calculation of the optical constant K.

a. Measure clear water.

b. Remove water and add molecular suspension.

c. Enter C of suspension in g/mL.

d. Measure suspension.

e. 'dn/dc' is automatically calculated & entered in Step 2 dn/dc box.

Molecular Weight - Sample Preparation for Debye Analysis

For each Debye plot MW measurement a set of samples are prepared covering a range of

concentrations. The range must be wide enough to be able to measure the slope

accurately (to determine the 2nd

Virial Coefficient), and the lowest concentration should

be close to the C=0 axis to minimize the extrapolation of the y-axis intercept (1/MW).

The units of C for the Debye plot are [(weight of molecules) / (total volume)],

(grams/ml). This is designated CWV. A set of 2 or 3 samples is sufficient to make a

Debye plot but up to five can be used. The concentrations must be accurate to

+/-1% in order to maintain a similar accuracy for the Molecular Weight and 2nd

Virial

coefficient.

Suggested set:

CWV1 = 0.002 - 0.005 gr/ml

CWV2 = 0.01 - 0.02 gr/ml

CWV3 = 0.03 - 0.05 gr/ml

Other units of concentration are used in sample preparation but the final samples must be

converted to CWV. The conversion requires the value of the molecular density, ρm, of the

molecule being measured. The other units of concentration are:

CVV = [(volume molecules) / (total sample volume)]

CWW = [(weight molecules) / (total sample weight)]

Conversion from one C to another requires the density of the molecules:

CWV = CWW / [ 1 - CWW(1 - 1/ρm) ]

CWV = CVV ρm

To prepare the set of concentrations it is most accurate to start by preparing the highest

concentration of the set first, and diluting portions of the high concentration mix to obtain

the lower concentrations.

OM0022 Operation


Preparation from Dry Powders:

The most accurate means of preparing the samples is to start with the molecular species

in the form of dry powder which can be weighed out to an accuracy of better then +/-1%.

A precision laboratory scale with +/- 0.1 mg reading error is recommended. The powder

is mixed with a solvent known to suspend the molecules. For example, Lysozyme

powder will dissolve in 0.1N sodium acetate water solution, and Bovine Albumin powder

into 0.1N NaCl water solution.

Example: Technique to prepare 20grams sample of CWW = 0.0200 gr/gr sample

* Place 25ml glass vial on scale, zero out tare.

* Weigh out powder in vial: Wsample = CWW*Wtotal = 20 x 0.02 = 0.4g of powder to an

accuracy of ±0.5% (±2mg).

* Calculate the exact total weight: Wsample / Wtotal = 0.02; Wtotal = Wsample x (1 / 0.02) =

50 x Wsample.

* Add water based solvent so the total weight = (50 x Wsample), to an accuracy of ±0.5%.

* The concentration is 0.0200 in (grams sample) / (grams total) = CWW

* Convert to CWV , in grams per ml: CWV = CWW / [ 1 - CWW(1 - 1/ρm) ]

Note: The density of the molecule is required to convert CWW to CWV.

Dilution from solution of known CWW:

If the solution has a known CWW it can be readily diluted using precise weighing.

* Start with a weight, W1,of known CWWHi.

* Add a measured weight of fluid, W2.

* Calculate CWWLo = [ (CWWHi x W1) / (W1 + W2) ].

* Convert to CWV, grams/ml. CWV = CWWLo / [ 1 - CWWLo(1 - 1/ρm) ]

Example: Technique to prepare approx. 20 grams of 0.002 grams/gram mix from 0.02

grams/gram mix

* Both weights together start as W1 + W2 ~ 20g

* CWWLo / CWWHi = 0.002 / 0.02 = 0.1. W1 = 20 x 0.1 = 2g of CWWHi mix needed.

* Add W2 = 20 – 2 = 18 grams of fluid to 2g of CWWHi mix.

* Convert to CWV , in grams per ml: CWV = CWW / [ 1 - CWW(1 - 1/ρm) ]

Dilution from solution of known CWV:

If the solution has a known and accurate CWV, it can be readily diluted using precise

volume pipette (volume measured to a precision of +/-1%).

* Start with a volume of V1 of CwvHi

* Add a volume of fluid V2 to the volume V1.

* Calculate lower concentration CwvLo = CwvHi x V1/(V1 + V2)

Example: Technique to prepare 20mL of 0.002 g/mL from 0.02 g/mL

* Both volumes together start as V1 + V2 = 20ml.

* CWVLo / CWVHi = 0.002 / 0.02 = 0.1. V1 = 20 x 0.1 = 2ml of CWVHi needed.

* Add V2 = 20 – 2 = 18mL of fluid to 2mL of CWVHi, to obtain 20mL of CWVLo.

OM0022 Operation


Molecular Weight - Measuring Molecular Density

The molecular density ρm can be determined using a density bottle and a sample mixed to

a known value of CWW.

A density bottle (such as Cole Parmer R-34579-20) has a known volume, Vo.

The weight of the bottle filled with a molecular solution of known CWW minus

the weight of the empty bottle yields the weight of the solution, Wo. The

solution density is calculated, ρs = Wo/Vo . For aqueous solutions CWW and

the solution density yield an expression for the molecular density:

WWs

sWWm

C

C

11

The molecular density is necessary to calculate CWV from CWW, and it is also

needed in order to dilute samples from high concentration to lower

concentration.

OM0022 Operation


3.5.6 Temperature Operation and SOPs

Additional information for using Microtrac Flex software to control the sample-cell

temperature can be found in Flex Software Manual SW0005.

CAUTION

Wave analyzers that are equipped with Temperature Control option have the capability of heating fluids to as high as +90°C. The user must take all precautions regarding heating of potentially hazardous fluids, including fluid flammability, ignition hazards, vapor hazards. etc.

When a Wave analyzer is chosen from the Measure menu, if it has the Temperature

Control option installed, then the Temperature toolbar will be displayed in the main

Measure Window:

The Temperature Control option allows control of sample temperature in 2 ways:

* 'Thermal Setpoint' control;

* 'Thermal Experiment' control;

NOTE: For Nano or Zeta models with Internal Probes, both Setpoint control and

Experiment control are available. For Nano models with External Probes,

only Setpoint control is available; Experiment mode is NOT available with

External Probe models.

Temperature - Setpoint Control

Click 'Tset' icon in toolbar to reach the Setpoint Control panel. This operation allows

simple entry of a desired temperature ('setpoint'):

OM0022 Operation


When the desired temperature is achieved, a message is presented:

The system will control at the temperature setpoint until the user stops the process, turns

the Temperature Control off, or enters another setpoint. During the time that temperature

is at the setpoint, the user can initiate a run, which will use parameters entered in sizing

and zeta SOPs to measure size (and zeta, if equipped and enabled).

Temperature - Temperature Experiment Control

Click 'Texp' icon in toolbar to reach the Experiment Control panel:

OM0022 Operation


The Thermal Experiment SOP allows entry of multiple temperature setpoints. The

Temperature Control system will automatically change to each temperature in the SOP,

stop, and run size and zeta (if equipped and enabled), using parameters entered in size

and zeta SOPs:

Temperature - Calibration of Temperature Control Hardware

The following section may not apply to all users of Wave with Temperature Control

Option. Instructions in this section requires correct combination of Flex PC software and

Wave internal firmware.

This section only pertains to Wave analyzers that have the Temperature Control Option

installed. This procedure shows how to use the Temperature Calibration function that is

in Flex, to improve the temperature-setpoint accuracy of the Temperature Control Option.

This calibration is intended to be performed at the final installation. The calibration

should then be able to account for conditions at the installation.

What Will Happen If This Calibration Is NOT Performed?

The Temperature Control Option of the Wave Analyzer will function even without this

additional calibration. If the calibration is not performed, the user may see a message

from Flex, each time that Flex is started and the Wave Analyzer is selected:

OM0022 Operation


This message may appear each time that Flex is started; once the calibration is

performed for the first time, then the message should not appear any more.

If the calibration is not performed, there may be some difference between a requested

Temperature Setpoint, and the measured sample temperature. For example, if the

requested Setpoint is 40°C, the actual measured sample temperature might be ~39°C.

The intention of the calibration is have the actual measured sample temperature to be

closer to the requested Setpoint Temperature, by accounting for the local conditions at the

installation site.

Performing the Calibration

The calibration takes about 1 hour and 45 minutes to perform and runs automatically

once it is started.

If Flex software is not already started, then start Flex, and connect to the Wave Analyzer

('connect' means to open a 'Measure' window within Flex).

Steps to perform the calibration are as follows:

1. Make sure the Wave Sample Cell has DI Water in it. Make sure that the Cell Cap is

installed in the Cell; the Cap will reduce the amount of sample evaporation that may

take place during the procedure.

2. In Flex software, go to Tools/Service/Utilities dialog screen.

3. In the dialog, click on the Temperature Tab that is along the top of the dialog.

4. Observe the “Temperature Set Points Calibration” section in the lower left.

OM0022 Operation


5. Click on the button.

6. Another screen will open and the calibration test will automatically start. The

calibration will collect data at 12 different Temperature Setpoints; this is automatic

and is not adjustable by the user.

Set Points Calibration Data Collection in Progress ( This example shows 'SetPoint 5', of 12 )

7. When the calibration procedure is complete the “Temperature Offset Table” (refer to

previous figures) will be filled with data.

OM0022 Operation


Completed Temperature Offset Table 8. IMPORTANT: When the calibration is complete, then in the Temperature Set Points

Calibration section, click the button to complete the calibration. This operation

does two things:

Sends the calibration factors to the Wave unit‟s RAM memory

Writes a file “Current.wtp” to the program folder where FLEX is installed.

The file (“Current.wtp”) is read into FLEX each time that Flex software is started and

then is connected to a Wave Analyzer. The file is read when a Wave Measurement

Window is opened, and then the data in the file is uploaded to the Wave RAM memory.

If this file is NOT found, then the start-up message (see the previous section of this

procedure) will be displayed.

Note: The contents in the “Current.wtp” file is not permanently stored in the Wave unit.

If power to the Wave unit is turned OFF, the values in the Wave RAM memory will

be lost. In this case, the Flex software must be closed, and then restarted, in order to

reload the calibration data into the Wave memory.

Optional Operations

Export the contents of the Temperature Offset Table to a disk file. As mentioned,

when the calibration procedure is complete, the data is stored in 'current.wtp'

file in the PC's folder for the Flex software. It is also a good idea to Export the

data to another file, with user-defined filename. This would be a backup file;

this would allow the user to Load a .wtp temperature calibration file at a later

time. For example, the 'current.wtp' file is automatically stored, but the user

could also Export to 'WaveTemperatureCalibration_July2012.wtp', or similar

filename.

Load the contents of the Temperature Offset Table from a previously exported disk

file.

OM0022 Operation


Print the contents of the Temperature Offset Table.

What If The Calibration Procedure Has a TimeOut On SetPoint 1?

For SetPoint 1 of 12 in the calibration procedure, the Wave analyzer system will

attempt to set the temperature to ~+3°C. Because there are no existing calibration

factors, the analyzer may not be able to reach the temperature in the set amount of

time. This could result in an error message, and the procedure would stop. If this

occurs, then close the Flex software, and cycle the power to the Wave ( switch OFF,

wait about one minute, then switch back ON ). Then restart the Flex software and re-

connect to the Wave analyzer by opening a 'Measure Window'.

The Temperature Control buttons should appear on the right-side of the Measure

window:

Click the 'Tset' button; the Temperature Setpoint screen should appear. Enter '5' for

the Setpoint, then click 'GoTo'; the analyzer should then try to set the temperature to

~+5°C.

The figure shown is for example only; the figure shows an increasing temperature

ramp; but for this step, the temperature should ramp down, from room temperature

down to about ~+5C.

When +5C is reached, the Setpoint screen will display a message.

Again, this figure is for example only; in this step, the temperature should be ~+5C.

OM0022 Operation


Click 'OK', then click 'Close' on the Temperature SetPoint screen. Then, go to Step 2

of the previous calibration procedure and try again.

OM0022 Operation


3.5.7 Titration Operation and SOPs

Additional information for using Microtrac Flex software to control the Zetrator titration

accessory can be found in Flex Software Manual SW0005.

Additional information for operating the Zetrator titration accessory can be found in

manual OM0014.

In order to use Zetrator titration accessory, Flow option must be installed on Wave

Analyzer (see previous section in this manual for the setup).

When the Zetrator titration accessory access is 'enabled' (see previous section in this

manual for setup), the 'autotitrator' button will appear in the toolbar of Flex

Measure Window:

Selecting the titrator button will launch the Autotitration Control Panel:

OM0022 Operation


All titration operations are conducted from this panel. Titration SOP parameters can be

set from this panel by selecting the 'SOP' button: . A titration experiment

results in a Data Display:

Further information about titration SOP parameters, titration calibration, selecting data

for display, printing reports, etc., can be found in manual OM0014.

OM0022 Operation


3.5.8 Combining SOPs

Some Wave SOP operation can be combined, but in limited fashion:

Siz

ing

Zet

a

Mole

cula

r

Wei

ght

Tem

per

ature

Set

poin

t

Tem

per

ature

Exper

imen

t

Tit

rati

on

Sizing

Zeta X X

Molecular Weight 1 X X

Temperature Setpoint X 2

Temperature Experiment X

Titration

1 For Molecular Weight operation, it is possible to use Temperature Setpoint to

set a temperature. However, Molecular Weight operation will then be

changing the sample in the Cell. This will make Temperature control of the

sample difficult. It is not recommended to combine Molecular Weight with

Temperature Control. 2 When connected to Zetrator, using Flow Cap option, it is possible to use

Temperature Setpoint to set a temperature. However, Titration operation will

then be changing the sample in the Cell, and the Cell is also connected to

Zetrator by tubing. This will make Temperature control of the sample

difficult. It is not recommended to combine Titration with Temperature

Control.

OM0022 Operation


3.6 Nanotrac Wave Testing Guidelines & Information

SetZero ('SZ')

SetZero is a procedure for measuring the background or steady-state noise of the system.

Some of this noise can originate from minute contamination that may be present in the

clean dispersing fluid, but the bulk of it is electrical noise inherent in the electronics of

the system. Noise characteristics can vary from unit to unit, but are inherently constant in

a given unit and are always present, whether clean fluid or a sample is being measured.

Therefore, it is legitimate to treat contamination and noise as a background. This

background is measured with clean fluid in the cell, stored, and then subtracted after each

sample measurement.

* Because some of the background noise can be due to conditions of the fluid or fluid-

probe interface, perform a SetZero whenever there have been changes in the environment

or operating conditions, such that the background may have changed. For instance,

SetZero must be performed whenever a new solvent or dispersing fluid is used.

* Perform a SetZero whenever the type of sample being measured is changed

dramatically (for example, changing from a mineral slurry to a paint emulsion).

* A SetZero must be performed after a power-up, before making any sample runs.

* A SetZero must be performed after opening a Nanotrac Wave measurement window in

the Microtrac® Flex Software.

* Before performing a SetZero, use a clean pipette (provided with system, or supplied by

the user), filled with appropriate fluid, to flush out the cell and remove all particles.

Fill the sample cell with clean dispersing fluid. Perform a SetZero using the Microtrac

FLEX Software Program.

CAUTION

Do not move, bump, touch or otherwise disturb the Nanotrac Wave during the SetZero procedure.

At the end of the SetZero procedure the computer screen will display the SetZero Status.

If the SetZero Status is BAD, then thoroughly clean and rinse the Sample Cell. Fill the

sample cell with clean fluid and use the Background function on the Microtrac FLEX

Software Program to check the Loading Index. The Loading Index should be less than or

equal to 0.010 to be acceptable to the system as background.

After flushing all particles from a cell, you can gauge its cleanliness by comparing the

measured Loading Index to the loading level currently being utilized. Flushing should

proceed until the loading level clean is 1/100 to 1/1000 the expected loading level with

current sample being run. For example if expected loading is 50 clean to 0.05 to 0.5, if

expected level is 2 clean to 0.002 to 0.02.

OM0022 Operation


Microtrac recommends that a SetZero be performed at startup each day. Then the

SetZero procedure should be periodically performed throughout the day. For optimum

performance, especially if using sample with low concentrations, Microtrac recommends

performing SetZero with clean liquid before every sample run.

Sample Dilution

Dilution is normally necessary because most processes provide fairly concentrated

samples. The sample acquired from the process must be diluted using a solvent or

dispersing fluid compatible with the particles and the Microtrac® system. The SetZero

measurement must be performed with the chosen fluid in the cell. When choosing the

fluid, its viscosity and refractive index must be known and entered into the fluid

parameter menu.

'Multiple Run' Selection

Multiple Runs may be chosen to provide multiple measurements and average

measurements of the same sample. Total time of Nanotrac Wave Multiple Run sequence

will depend on:

* The „Run Time‟ that is entered by user;

* If Zeta function is available, is it Enabled; total zeta run-time will somewhat depend

on the sample itself.

If Zeta function is Enabled, a Nanotrac Wave measurement run consists of 1) a sizing

measurement, 2) and zetapotential magnitude measurement, and 3) a zetapotential sign

measurement. If „n‟ Multiple Runs are chosen, the overall sequence is as follows:

1) Size measurement 1; time = Run Time;

2) Zeta magnitude measurement 1; time = Run Time;

3) Zeta sign measurement; time = minimum 20 seconds, maximum possible time

depends on sample;

4) Size measurement 2; time = Run Time;

5) Zeta magnitude measurement 2; time = Run Time;

.

.

.

*) Size measurement „n‟; time = Run Time;

*) Zeta magnitude measurement „n‟; time = Run Time;

Multiple Measurements of the Same Sample

If Zeta function is available and Enabled, then the Nanotrac Wave‟s zetapotential

measurement operation has the possibility of altering the particle distribution throughout

the sample's volume. The possibility that this alteration will happen is sample-dependent.

Therefore, Microtrac recommends that if repeated measurements are to be made of the

same sample, with „Zetapotential‟ enabled, that in-between measurements the sample

should be agitated to redistribute particles throughout the sample volume. For instance,

using a clean micropipette, the sample could be extracted and reinserted several times;

this will perform the redistribution action.

Note: this does not imply that any user action is needed during „Multi Run‟

measurements; agitation action may only be necessary after the entire run is complete.

OM0022 Operation


Fluid Compatibility

The Nanotrac Wave has been designed for use with a wide range of fluids. However,

when using non-aqueous samples (polar solvents, non-polar solvents, organic solvents,

pH outside of stated range, etc.), the user is recommended to contact Microtrac Technical

Support. Refer to Specifications section for more information.

Fluid Viscosity

Fluid viscosity must be known accurately at two temperatures within the +5o to +90

oC

operating range. The operator enters these two temperatures and corresponding viscosity

values into the Microtrac® FLEX software. FLEX uses these values and the measured

cell temperature to compute actual fluid viscosity at the measurement temperature.

Viscosities from 0.3 to 3 cp are generally preferred. Higher viscosities cause slower

particle velocities and, therefore, lower frequencies in the detected signal. The upper size

limit will be determined by the viscosity -- the higher the viscosity the lower the upper

size limit. In water (1cp) the size limit is 6.4 microns. At a viscosity of 10cp the limit

would be 0.64 microns.

A signal with a frequency spectrum mainly in the low frequency range requires longer

sampling time, with the time increase proportional to the viscosity increase above that of

water. For example:

* A 500 nm sample has normal run time of 30 to 60 seconds in water, 1 cp fluid

viscosity.

* A 500 nm sample in a fluid of 3 cp viscosity exhibits a frequency spectrum similar to

that of a 1,500 nanometer sample in water. Thus, a 500 nm sample in a fluid of 3 cp

viscosity must use a run time of 90 to 180 seconds or more.

* The Nanotrac Wave has a particle size range of 0.0008 (0.8nm) to 6.54 microns in

water. Higher viscosity changes the upper size limit. The effect can be estimated as

follows:

- The product of [ Viscosity (cp) * Particle Diameter (microns)] should be between

0.0008 (0.8nm) and 6.54;

- A 1 micron sample in 3 cp fluid is equivalent to a 3 micron sample in water (1 cp);

- A 2 micron sample in 3 cp fluid is equivalent to a 6 micron sample in water.

Refractive Index

Particles are visible to the instrument only if their refractive index is different from that

of the suspending fluid. Scattered power is reduced as the particle index approaches the

fluid refractive index. Refer to SW0005 Flex Software Operations Manual for additional

guidance on how to enter refractive index information for fluids and for particles. Use

the SAMPLE LOADING function and the procedures outlined in the next section, to

determine if the fluid/particle combination gives adequate Loading Index (LI).

Sample Concentration

The Loading Index is given at the end of a measurement and in the Sample Loading

window; refer to SW0005 Flex Software Operations Manual for details. The Loading

Index is a measure of the total AC signal obtained from the light scattered from the

moving particles.

OM0022 Operation


Particles with a high refractive index relative to the fluid index tend to give larger signals.

However, there can be significant variation of scattering efficiency as a function of

particle size. Efficient scatterers can give adequate signal at concentrations as low as a

few parts per million (ppm), while inefficient scatterers can require concentrations of

several thousand ppm.

Scattering efficiency makes concentration important for some samples. Overloaded

samples can create optical signals which could create artificial „modes‟ or peaks in the

size distribution. Scattering efficiency needs to be considered when sample concentration

is determined.

Concentration limits can be broken down into three ranges depending on the Loading

Index.

Safe Range: Loading Index = 0.1 to 100.0

This range of Loading Index can be used without concern for instrument or sample

limitations.

Caution Range: Loading Index < 0.1

Use very small particle or very low concentrations of particles with caution. There is no

absolute lower-Loading-Index limit, but the precautions observed become more severe

depending on the Loading Index utilized. Concerns include:

Contamination of small particles with low concentrations of large particles.

SetZero Background Index should be less than 0.010

Interference of the measurement with environmental changes, such as bench vibration

or rapid temperature changes should be avoided.

Caution Range: Loading Index > 100.0

High Loading Index values are also to be used with caution. At high sample

concentrations, particle interactions become more probable, and results are subject to

interpretation. Agglomeration, changes in the effective dispersing fluid viscosity, and

changes in effective dispersing fluid index can result from high sample concentration and

particle interactions. These effects can shift or distort measured distributions. The

individual sample chemistry will determine the Loading Index limit. Determination of

the limits must be made on a sample-by-sample basis.

3.7 Sample Preparation

Some applications require the measurement of particles that are normally dispersed.

These samples require no preparation and can be added directly to the Sample Cell.

Other applications require the addition of surfactants or other dispersing agents, in

addition to mechanical energy, to disperse the sample into individual discrete particles.

Samples must be representative of the entire product lot or batch. This determines which,

if any, sample preparation techniques are required. Sample preparation techniques

presented in this section are of a general nature; address specific questions about

particular sample preparation techniques to:



Largo, FL 33773

(727) 507-9770

OM0022 Operation


Sample Preparation Techniques

Two steps, wetting and dispersion, are normally required to achieve a non-agglomerated

sample.

* Wetting uses water, other suspending media, and/or chemicals to reduce surface tension

to promote mixing and diluting a sample in a suspending fluid. A wetted sample mixes

freely.

* Final dispersion (de-agglomeration) may require energy to act mechanically on the

suspension.

Wetting agents include water, surfactants, dispersants, and solvents.

NOTE: Surfactants or dispersants used in excess can cause formation of bubbles that

may affect the measurement. Excess surfactant can also lead to re-agglomeration when

reaching the surfactant critical micelle concentration.

NANOTRAC WAVE NOTE: Use of surfactants and dispersants may affect the accuracy

of Zeta measurements.

Dispersion

Dispersion requires energy input from one of several devices. Most commonly,

ultrasonic energy is employed in the form of a bath or probe.

High shear devices such as tissue homogenizers should not be used because they tend to

produce artificial distributions and a non-representative sample.

Germicides

Microscopic organisms in the sample or the dispersing fluid can be read as particles. Use

an appropriate germicide if the fluid is conducive to microbial growth. Such growth is

typically slow. In clean water, bacteria can take several weeks to grow to a noticeable

size and/or concentration.

Particle Size Measurement

Once the sample has been diluted to a concentration between the upper and lower

boundaries of the loading screen, it can be run. Follow the instructions above, and in the

SW0005 Microtrac FLEX Software Manual, to conduct a run.

Run Time

Optimum measurement (run) time depends on the particle size. Small, fast-moving

particles can be measured in short times, while larger slower moving particles require

longer times. Use the following guidelines to determine minimum measurement time.

Measurement times needed to achieve maximum repeatability are determined

experimentally by calculating statistical repeatability (for example mean and standard

deviation) at several measurement periods.

Table 1-1 Run Time

Particle Size Range

(Nanometers)

Minimum Run Time

(Seconds)

Below 60 30

60 to 300 90

300 to 900 120

Above 900 180

OM0022 Operation


Run time can always be increased above these values. Longer Run Times may provide

more repeatable data.

After completing a sample measurement, thoroughly flush the sample cell to avoid

accumulation of contaminating particles. Refer to the Maintenance section for additional

guidance on cleaning the sample-cell. Do not leave sample material in the cell. The fluid

can evaporate and deposit particles on the probe face and other cell parts. Dried particles

are more difficult to remove than suspended particles.

If particles dry onto the sample cell surfaces, wet-swab the entire sample cell. Then rinse

the sample cell two to three times with clean fluid to remove any residual contaminates.

The Sample-Cell can also be removed for further cleaning, such as by ultrasonic bath,

sterilization, etc. See other sections in this manual for more information.

OM0022 Principles of Operation


4. Principles of Operation

4.1 Introduction

The particle-size measurement technique used in Nanotrac Wave is that of dynamic light

scattering (DLS). The velocity distribution of a sample of particles suspended in a

dispersing medium is a known function of particle size. Light from a laser diode is

coupled to the sample through an optical power splitter/probe assembly. Light scattered

from each particle is Doppler-shifted by the particle motion (Brownian motion). The

Doppler-shifted scattered light is mixed with coherent, un-shifted light; and the optical

system sends these mixed signals to a silicon photo-detector. The detector output signal

is then amplified, filtered, digitized, and mathematically analyzed by the Microtrac®

FLEX Windows Software, using proprietary algorithms, to provide the particle size

distribution.

Nanotrac Wave extends the dynamic light scattering measurement function of Microtrac's

Nanotrac™ particle size analyzer, to measure the particle-size-distribution, the molecular

weight, and, when equipped with zeta option, the zetapotential of suspended particles.

Added to Nanotrac™ is the ability to apply an electric field to a particle suspension and

simultaneously view the resulting particle motion. Analysis of the particle motion

determines the particle charge, electrophoretic mobility and zeta potential. In addition to

the zeta analysis all measurements include the standard Nanotrac™ particle size

distribution.

4.2 Physical Principles

Brownian Motion

Particles suspended in a dispersing fluid are subject to random collisions with thermally

excited molecules of the fluid. The velocity and direction of the resulting motion are

random, but the velocity distribution of a large number of mono-sized particles averaged

over a long time will approach a known functional form.

The velocity distribution is a known function of particle size. Figure 23 shows that a

sample of small particles has a higher median velocity than a sample of large particles,

where the median velocity is inversely proportional to the particle size. Velocity

distribution is also a function of fluid temperature and fluid viscosity.

If the fluid molecules have higher average thermal energy (higher temperature), they will

impart higher velocities to the particles with which they collide. Median particle velocity

is directly proportional to the absolute (Kelvin) temperature of the fluid. A viscous fluid

slows the energized particles. Particle velocity is inversely proportional to fluid

viscosity.

Viscosity itself is a complex but predictable function of temperature. If the operating

temperature is known, the viscosity at that temperature can be computed. Thus,

compensation can be made for both temperature and viscosity effects. With

compensation, the velocity distribution becomes a unique function of particle diameter.

Viscosity entered into the Flex software must be of the clean fluid only; considerations

of the viscosity of the mixed sample, especially if that mix is considered to have non-

Newtonian behavior, must be avoided.



Figure 24 shows the general behavior of particle velocity distributions as a function of

particle diameter, fluid temperature, and fluid viscosity. The median particle velocities

range from 5 micron/second to 6000 micron/second.

Doppler Effect

Light from the laser is coupled through one of the fiber optic tails, through the sapphire

window and into the fluid. A small percentage of the light is reflected from the

sapphire/fluid interface and travels back through the fiber and into the photo-detector

mounted on the Laser/Detector board. Doppler-shifted light scattered back from the

particles enters the probe at the tip, mixes with the reflected light and travels back to the

photo-detector.

Light incident on a particle scatters in all directions. If the particle is stationary the

scattered light is of the same frequency (or wavelength) as the incident light. If the

particle is moving at some velocity relative to the light source, the scattered light is

shifted in frequency by an amount proportional to the particle velocity. An ensemble of

particles with a certain velocity distribution will thus have a unique distribution of

frequency shifts.

2 micron particles

0.005 micron particles

Velocity , microns/second

N

N/2

V1 V2

a/n 23142

Figure 37: Velocity Distribution, Suspended Particles in Brownian Motion



2 micron particles


N

50°C Water

(or low v iscosity liquid)

2 micron particles


N

10°C Water

(or v iscous f luid - i.e. isopropy l alcohol)

2 micron particles


Velocity

N

N/2

Room Temperature Water

a/n 23143

Figure 38: Velocity Distribution, Temperature and Velocity Effects



Heterodyne Detection

Heterodyne detection involves combining Doppler (frequency) shifted light scattered

from moving particles with a reference beam of light from the same source, not Doppler

shifted, reflected from a stationary surface; see Figure 25.

The particle velocities are so small compared to the velocity of light that the Doppler

frequency shifts can only be detected using frequency-beating techniques. Light reaching

the photo-detector consists primarily of a large amplitude component at the transmitted

frequency, Ft, and much smaller amplitude components at the frequency of the Doppler-

shifted scattered light, F1 and F2.

Because the laser output and the reflection at the probe sapphire/fluid interface are

constant, the reflected component, Ft, will be large and constant. The components F1 and

F2 will be small and varying as the particles continuously change direction and velocity.

Thus the detector will generate:

* A large dc output component proportional to the reflected light

* Smaller ac output components at the heterodyne difference frequencies, (F1 - Ft) and

(Ft - F2)

* Much smaller ac output components at the self-beating frequency, (F1 - F2)

Figure 39: Heterodyne Detection of Scattered Light

The self-beating or homodyne frequencies are twice the value of the heterodyned

frequencies and can be considered interference. The Nanotrac Wave Analyzer makes use



of the heterodyned signal, which will be the major signal over a very wide range of

particle concentrations.

Frequency Spectrum

In a real measurement situation, large numbers of particles move randomly in the vicinity

of the probe tip, generating an equal number of Doppler-shifted scattered light signals.

These signals of various frequencies combine with the reflected signal of un-shifted

frequency to generate a wide spectrum of heterodyne difference frequencies. The

resulting output of the photo-detector is thus a random signal with a frequency spectrum

determined by the velocity distribution of the particles in the sample.

The random signal from the photo-detector is sampled and digitized by the analog-to-

digital converter board (data acquisition board). The resulting stream of numbers then

undergoes a complex series of mathematical operations designed to compute the

frequency spectrum of the original random signal.

Frequency shift is directly proportional to particle velocity, so the shape of the frequency

spectrum as a function of particle size, temperature, and viscosity is very similar to the

velocity distributions shown in Figure 23. Spectra from small particles have more high

frequency components than those from large particles. A higher fluid temperature (or a

lower viscosity) will cause increased high frequency components in the spectrum.

Particle Distribution

The frequency spectrum is uniquely determined by the particle velocity distribution that,

in turn, is uniquely determined by the particle size distribution. Using proprietary signal-

processing algorithms, the particle distribution is computed directly from the measured

frequency spectrum recovered from the Doppler-shifted scattered light.

Interference Effects and Scattering Efficiency

Transparent particles give rise to optical interference effects; many particles in the

Nanotrac Wave size range are transparent. These effects can be visualized by reference

to Figure 25.

What is not shown is that a portion of the incident light, at the Doppler shifted frequency

F1, enters the particle and travels inside the particle material at a velocity determined by

the index of refraction of the particle material. After traveling through the particle, the

light hits the "far wall" and some of it "bounces" and travels back through the particle. A

portion of this bounced light then exits at the front wall and travels toward the probe on

the same path as the light scattered from the front wall. The frequency of both waves is

the same but the phase is not. Since the bounced wave has been delayed by traveling

back and forth through the particle, its' phase fronts (or minima and maxima) cannot align

exactly with the phase fronts of the scattered wave. The difference in the alignment of

phase fronts is called optical interference, which can be constructive or destructive:

In the case of constructive interference, the phase fronts align exactly:

* The combined wave has a higher amplitude;

* The particle is an efficient scatterer;



In the case of destructive interference the Maxima of one wave align with minima of the

other wave:

* The combined wave has a lower amplitude;

* The particle is an inefficient scatterer

The amplitude of the signal recovered at the photo-detector will be larger or smaller

depending on whether the particles cause constructive or destructive interference. The

size of the particle and its refractive index will determine the delay between the two

waves and therefore the nature of the interference and scattering efficiency.

Figure 26 is a plot of scattering efficiency versus particle size, for a given combination of

particle index and fluid index. Figure 27 gives an expanded view of the scattering

efficiency in the region of 0.0 to 0.1 micron.

Other fluid/particle combinations will yield a similar plot but the peaks and valleys will

be at different values of particle size.

If the particle distribution in a sample consists of a single particle size, optical

interference is not a problem. The shape of the frequency spectrum is unaffected,

resulting in the proper computation of particle size.

Optical interference effects can be significant if the particle distribution in a sample is

bimodal, or a broad distribution of sizes. The computed particle size distribution would

be distorted, skewed in favor of the more efficient scatterers. Note that in the Rayleigh

region (up to about 200 nm) the change in scattering efficiency varies rapidly with

particle size.

When the distorting effects of the scattering efficiency function are properly

compensated, the computed particle size distribution is a true Volume Distribution. If the

distorting effects of the scattering efficiency function are uncompensated, the resulting

computed particle size distribution is more properly termed an intensity Weighted

distribution.

Instruments based on older technologies were unable to make the necessary

compensation and presented only Intensity-Weighted particle size distributions.

Microtrac® Particle Size Analyzers compute true Volume Distributions in the standard

mode of operation, and offer the user the alternative of selecting Intensity-Weighted or

Mono-Disperse modes of operation. Refer to SW0005 Flex Operations Manual for

details on how to selection the mode of operation.

Norm

aliz

ed S

ignal

Signal per Unit Volume v s. Size

Poly sty rene in Water

0.0 0.2 0.4 0.6 0.8 1.0 1.2 1.4 1.6 1.8 2.0 2.2 2.4 2.60.1 0.3 0.5 0.7 0.9 1.1 1.3 1.5 1.7 1.9 2.1 2.3 2.5 2.7

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0

Size (micron)

a/n 23145

Figure 40: Scattering Efficiency, Full Range



Norm

aliz

ed S

ignal

Signal per Unit Volume v s. Size

Poly sty rene in Water

0.0 0.01 0.02 0.03 0.04 0.05 0.06 0.07 0.08 0.09 0.100.0

0.10

0.15

0.20

Size (micron)

0.05

a/n 23146

Figure 41: Scattering Efficiency, Rayleigh Range

4.3 Zetapotential Measurement

As discussed in previous sections, the Nanotrac Wave has all of the features of

Nanotrac™, with additional zetapotential measurement capability provided by new

hardware:

1. A voltage source with programmable amplitude and wave form.

2. An insulating sample cell, with optical probes opposed by electrodes, forming cell

which makes zetapotential measurement possible.

3. Optical probes, with sample interface window consisting of typical sapphire, but

having specialized metallic and semiconductor optical coatings applied.

4. Coated-window optical probes are paired with their opposite electrodes. Excitation of

cell, from the above voltage source, applied between the optical probe and it's electrode,

creates electric fields. Particle motion is analyzed while under the influence of the fields.

As mentioned previously, particle size distribution is determined from the velocity

distribution of particles suspended in a dispersing medium, using the principles of

dynamic light scattering. The Nanotrac Wave analyzer measures the additional velocity

imparted to the charged particles when placed in an electric field. Particle electrophoretic

mobility is calculated from this additional velocity component. Zeta potential is

calculated from mobility using accepted relationships between mobility and zeta

potential. In the limit of high concentration of electrolyte, the relation between zeta

potential and mobility is given by the Smoluchowski equation:

= zeta potential, = mobility, = viscosity, = dielectric constant

For water at 25degC, Zeta potential(mV) = 12.8 x Mobility(μ/sec/volt/cm)



4.4 Molecular Weight Measurement

Prior to the 10.6.1 version of Flex software, a calculation of a sample's Molecular Weight

was calculated and displayed in the Summary table. The calculation was based on

measured hydrodynamic properties.

The accuracy of this estimate is dependent on the particle's characteristics, and how close

these characteristics are to true molecular hydrodynamic behavior.

Beginning with release 10.6.1 of Flex, Microtrac included another Molecular Weight

calculation feature, based on a measurement of molecular suspension scattered light

intensity, and utilizing the well-known Debye Plot technique.

Debye Plot and It's Molecular Weight Calculations

The measurement of Molecular Weight ('MW') using the Debye plot depends upon an

accurate measurement of the light that is scattered by a molecular suspension of known

concentration, C. The ratio of total scattered-light to total incident-light is the Rayleigh

ratio, R. The Debye MW expression relates the MW to the Rayleigh ratio and to the

sample concentration:

CAMR

KC

W22

1 (1)



This relationship has the linear form of y = b + mx, and can be plotted:

where:

C is the Sample Concentration in units of grams/ml

R is the Rayleigh ratio of scattered-light to incident-light at any concentration C

K is an optical constant dependent on the molecule, solvent, and incident light:

2

4

24

dc

dnn

NK o

Ao

(2)

NA = Avogadro's number 6.022 x 1023

molecules/mol

λo = wavelength of incident light. For Microtrac DLS analyzers, 780nm

no = refractive index of suspending medium. For aqueous sample, 1.333.

dc

dn = differential refractive index.

For many mixtures of molecular samples and solvents the value of dn/dC is

found in published literature. For unique combinations Microtrac DLS

analyzers, run with Microtrac Flex 10.6.1 software, can measure this value

in situ. See other sections in this manual for more information.

KC/R at C=0 is the y-axis intercept equal to WM

1. The inverse of this value is

Molecular Weight.

2A2 is the slope of the plot. A2 is the 2ND

Virial Coefficient, which is a measure of

sample/solvent interaction. If the coefficient (slope) is positive, the sample tends to

stay dispersed in solution as concentration is increased. If the coefficient is negative,

the sample tends to agglomerate as concentration is increased.

Debye Molecular Weight With Microtrac Dynamic Light Scattering Analyzers

For Microtrac DLS Analyzers, the angle of incident-to-scattered light is 180°

(backscatter). The Rayleigh Ratio is calculated from the DLS Analyzer's measurement of

the backscattered light intensity. The backscattered light intensity is only a portion of the



total scattered light constituting the Rayleigh Ratio. The measured backscattered light

intensity, multiplied times a constant, equals the Rayleigh Ratio that is used. The value

of the constant is dependent on the fixed optical geometry of the DLS Analyzer's optical

probe, and is the same from unit to unit. The constant has been determined from back

scattering measurements on samples of known molecular weight.

When measuring molecular scattering there is often interfering scatter from small

concentrations of larger particles present. Larger particles have larger scattering

coefficients, due to inherent dependence of Rayleigh scattering intensity with particle

diameter D. Further, this dependence is non-linear, as scattered intensity increases as D3.

Even at low concentrations large particles can have scatter intensity comparable to the

smaller molecules of the molecular suspension. (For example: 1ppm / 100nm

contaminating a 10,000ppm / 5nm molecular suspension would have equal scattering

intensity and cause a 2 times error in the Molecular Weight). Microtrac‟s unique

Molecular Weight Analysis separates the molecular-suspension scattering intensity from

interfering larger-particle scattering intensity, by utilizing the Brownian motion signature

measured with the Analyzer's dynamic light scattering techniques. The ability to separate

the interfering scatter avoids complex mechanical filtering of molecular suspensions.

Molecular Weight By Hydrodynamic Sample Properties

The size of suspended molecules is determined by Dynamic Light Scattering technology,

the foundation of the Microtrac DLS Analyzer measurement. The DLS technology

measures the Brownian motion of the molecules and determines the molecular size

through the application of the Brownian motion principles. The size is referred to as the

hydrodynamic size. To calculate the Molecular Weight from hydro size, the molecule is

assumed spherical with a volume and weight of:

6

3Dvm

(3)

mmm vw (4)

where:

m = sample's molecular density (in Flex software, this is entered as part of SOP data);

Molecular Weight is grams per Mole of the molecule:

mAwNMW (5)

Therefore Molecular Weight is calculated from the molecular hydrodynamic diameter

and molecular density:

6

3DNvNwNMW mAmmAmA

(6)

OM0022 Maintenance


5. Maintenance

5.1 Introduction

This chapter contains the following information:

Routine Maintenance

Troubleshooting

Safe Maintenance Practices

5.2 Safe Maintenance Practices

Observe all safety notices posted on the Nanotrac Wave and throughout this manual.

WARNING

The Nanotrac Wave must be serviced by a qualified and authorized service technician. Do not attempt service procedures not described in this manual. Contact your Microtrac representative if questions arise concerning operation, adjustment, or repair of any particle size analyzer.

5.3 Routine Maintenance

A program of regular inspection and maintenance helps to ensure continued optimum

performance of the Nanotrac Wave Particle Size Analyzer. The user can perform the

routine maintenance procedures in this chapter at intervals that are determined by the

amount and severity of use of the Nanotrac Wave.

Routine maintenance involves cleaning the sample-cell, and the optical probes.

5.3.1 Cleaning of Surfaces

The Nanotrac Wave has been built with tough, chemically resistant coatings, to provide a

good appearance for years of normal use. If the top surfaces of the Wave become dirty,

or some liquid is spilled on it, a clean, damp cloth can be used to wipe the surface. If

necessary, a mild solvent such as isopropyl alcohol can be used to try to remove stains.

Use of abrasive cleansers is not recommended as this will likely scratch the coated

surfaces.

5.3.2 Cleaning of Nano or Zeta Sample-Cells and Optical Probes Without Removing the Cell

To clean the Sample-Cell and Probes on Nano and Zeta models, without removing the

Cell:

1. First thoroughly flush out all particles from the cell. The user should be especially

careful if the particles are abrasive; in these cases, do not wipe the optical probe tips

until the cell is generously flushed.

2. Wipe the probe tip gently to loosen accumulated material. Supplied with Nanotrac

Wave are either *'Microbrushes', *Chamois swabs, or both. Moisten brushes or

swabs with deionized water.

See the figures that follow. For Zeta Cells, note that the chamois swabs have a cut-

out area on one side; insure that this side faces the 'electrode'. The swab will then fit

!

OM0022 Maintenance


between the electrode and the face of the optical probe, allowing for gentle cleaning

of the probe. Use up-and-down motion, applying gentle pressure to the probe face.

When using microbrushes, again, apply gentle pressure to the face of the optical

probe.

Clean each optical probe individually using these techniques.

Swabs and microbrushes are re-usable. Microtrac recommends using one swab or

brush for each type of sample to be tested, and to regularly use new swabs/brushes,

and dispose of used ones.

Flush the cell again several times to remove loosened material. The FLEX software

can be used to judge the cell cleanliness; use the 'Background BKG' or 'Loading LD'

toolbar buttons to activate a loading screen; these displays can be used as diagnostic

aides to determine the quantity of particulate in the cell. See previous sections of this

manual, and refer to Flex Operating Manual SW0005, for further use of these

functions.

3. Other solvents may be more appropriate for cleaning, depending on the nature of the

particles or contaminating materials. The user should consult Microtrac® Technical

Support for use of other solvents.

Close-up view of chamois swab

5.3.3 Cleaning of Sample-Cells and Optical Probes By Removing the Cell

If needed, the Sample-Cell can be removed for further cleaning.

Note: it is not necessary to turn off power to perform this cleaning, but the user should

follow all safety precautions in this manual.

OM0022 Maintenance


Cleaning of Sample-Cells and Optical Probes By Removing the Cell (continued)

First, open the Access Cover.

Figure 42: Gaining Access to the Sample-Cell Area

Figure 43: The Sample-Cell Area Under the Access Cover

OM0022 Maintenance



With the Cover raised, now the Cap is lifted away from the Cell:

1: Vent the Cap 2: Raise the Cap Lifter 3: Rotate the Lifter

Figure 44: Moving the Cap Out of the Way

CAUTION

Wave analyzers that are equipped with Temperature Control option have the capability of heating fluids to as high as +90°C. Users should always allow hazardous fluids to return to room temperature before venting or removing the Cell Cap.

When the Cover is removed, the Sample-Cell, the Cell Holder, the Probes, and the

Electrodes are exposed:

Figure 45: Remove the Cell Support Cover (Blue) Top: Zeta cell Bottom: Nano cell

OM0022 Maintenance



WARNING

Wave analyzers that are equipped with Temperature Control option have surfaces that can reach high temperatures. If the 'Hot Surface' symbol is present on the LCD display, AND if it is blinking, then a burn hazard is imminent and extra precaution should be followed. Users should always allow the Cell to return to room temperature before removing the Cell Support Cover.

Further description of the Cell Support assembly, including Sample-Cell, Cell-Holder,

Probes, Probe-Holder (Zeta model is the example shown).

Figure 46: Description of Cell Support Assembly

Additional descriptions of these components for both Zeta and Nano models is shown in

the following figures.

Both for Nano and for Zeta models, it is important to note the orientation of Probes,

Electrodes, and Plugs. When the components of the Cell-Support are disassembled for

cleaning, they MUST be re-assembled in the original positions for correct operation.

!

OM0022 Maintenance



Figure 47: Cell Support Components, Nano Model. Very Important: Note the Orientations

Figure 48: Cell Support Components, Zeta Model. Very Important: Note the Orientations

OM0022 Maintenance



Using the Wrench Tool that is provided with Wave analyzer, slightly loosen 4 screws on

the Probe Clamps:

Figure 49: Loosen Probe Clamp Screws

Then use the Probe-Holders to pull the probe(s) out of the Sample-Cell (if zeta model,

electrodes will also be pulled out of the Cell by the Holder):

Figure 50: Remove Probe(s) From the Cell

OM0022 Maintenance



Figure 51: Remove Probe(s) From the Cell

When the Probe(s) are free from the Cell, the Cell can be removed from the Cell-Holder;

the Cell can then be ultrasonically cleaned, sterilized, washed, etc.:

Figure 52: Remove Cell for Cleaning

OM0022 Maintenance



By using the Probe-Holders, the Probe(s) (and also Electrodes, if it is zeta model) can be

further removed from the Probe Clamps, and the tips of Probes and Electrodes can also

be cleaned. Use cleaning kits, swabs, and cleaning agents that are supplied by Microtrac.

WARNING

Laser-radiation is emitted from the apertures of the Optical Probe(s) at <1mW optical power levels. These apertures are accessible to the user of the Wave. All laser-safety precautions should be followed when handling Wave Optical Probes.

To re-assemble the Cell, first, using the Probe-Holders, gently insert Probe and

Electrodes back into the Probe Clamps.

Remember to re-assemble with the original orientations and positions.

Insert the Sample-Cell back into the Cell-Holder. The Cell is symmetrical and it is not

polarized - it can go into the Cell-Holder in either direction.

Then use the 'Gap' Tool that is provided with the Wave analyzer; one end of the Tool is

for Nano Cell, the other end of the Tool is for Zeta Cell:

Figure 53: Reassembly of The Cell, and The 'Gap' Tool

OM0022 Maintenance



* Insert the appropriate end of the Gap Tool into the Sample-Cell until it is seated

against it's stops; the example that is shown is for zeta Cell.

* Use Probe Holder to push Probe(s) and Electrodes into the Cell, until they contact the

Gap Tool.

* Remove the Gap Tool from the Cell.

* Again push Probes and Electrodes just a little further into the Cell, being careful to not

make contact with probe or electrode that is on the opposite side.

* Re-insert the Gap Tool; this will force the Probes and Electrodes back outward, into

their final position.

* Use the provided Wrench Tool to re-tighten four Probe Clamp screws. Do not over-

tighten.

* Again remove the Gap Tool from the Cell.

Figure 54: Use the Gap Tool to Re-set The Positions of Probe(s) and Electrodes

Finally, replace the blue Cell Support Cover over the Cell; then turn and lower the Cap

Lifter so that the Cap is inserted back into the Cell; then close the Access Cover. The

analyzer is ready to test the next sample.

OM0022 Maintenance


5.4 Troubleshooting

The need for troubleshooting arises under the following conditions:

Warning or error messages occur during operation.

Spurious or inconsistent data.

Should these situations occur, the user may be able to troubleshoot and fix the problem.

Before contacting Microtrac, the user should follow the steps below.

Troubleshooting: Is The Nanotrac Wave Properly Connected, And Does It Power-Up Correctly?

Refer to previous sections on installation and connection; check that all connections are

correct, and that connectors are fully inserted. Check for any visible damage to any

connectors (bent connector, damaged connector housing, etc.). Note: The user should

avoid connecting or disconnecting any Nanotrac Wave connections or computer

connections without first powering down the system. If a connector is found to be loose

or disconnected, first power down the system, then reconnect the connector, then power

up the system again.

Check that the AC power source is with specification, both for Nanotrac Wave DC power

supply and for computer.

Observe the Nanotrac Wave LCD display:

* With DC power supply plugged in, and power-switch turned on, is there any

activity on the display?

* On opening of Flex software, does Flex 'connect' to the analyzer, or is there an error

message, such as 'Failed To Find a Wave Device'? Contact Microtrac® service

for more information.

Troubleshooting: What Are Results of 'Bench Status' Check?

Nanotrac Wave performance can be checked with Flex 'Bench Status' diagnostic

function..

Prior to running Bench Status, if necessary, flush and clean the sample-cell as described

in Maintenance section. Fill the sample-cell with clean fluid. Click 'Tools - Service -

Bench Status' as shown below. Note: A Nanotrac Wave 'Measurement' window must be

open, and must be the active window, as shown below, in order to access the Bench

Status function.

Accessing Bench Status Function From An Active Measurement Screen

OM0022 Maintenance


Checked / Retrieved Parameters:

The following items are checked or retrieved during a Bench Status update:

Laser operation;

Data acquisition hardware;

Temperature Control hardware, if equipped;

The present status of the system, such as Access Cover interlock.

Tested Optical Parameters:

Reflected Power

Optical probe cleanliness, or other probe issues, with affect „reflected power‟; this may

lead to a Bench Status Reflected Power status of BAD.

Clean the Nanotrac Wave probe face(s) and fill the sample cell with clean fluid. Repeat

the bench status. If the Reflected Power remains high, an electrical alignment or other

action may be required. Call Microtrac® service for more information. Possible causes

are:

Probe disconnected

Fiber connection at laser not seated all the way

Bad or broken probe optical fibers

Defective/misadjusted Laser Drive Circuit

Defective Detector

Laser status

If the laser(s) are not operating properly then the status is BAD. If the status indicates

BAD, contact Microtrac® Service for more information. Possible causes are:

Ambient operating temperature too high

Laser drive circuit misadjusted

Defective laser driver device

Defective laser

Troubleshooting: Do Reference Samples Test Correctly?

Microtrac makes available particle size reference kits to assist in performance evaluation.

These kits contain samples of known properties. In cases where the performance of the

Nanotrac Wave is in question, a sample of reference material may be analyzed, and, if

desired, the results sent to Microtrac for evaluation.

Instructions for proper use of the reference material and expected results are provided

with the kit. Do not test with materials from an expired kit; expiration dates are printed

on the outside of the kit.

Some reference samples are provided with the Nanotrac Wave system. Contact your

representative or Microtrac to determined which reference materials are available, and to

purchase additional reference materials.

OM0022 Maintenance


Troubleshooting: Are Warning or Error Messages Displayed During SetZero or Run?

During a measurement operation, for example RUN or SETZERO, the Nanotrac Wave

system continuously monitors various critical parameters such as laser power level and

total reflected power. If any parameters fall outside acceptable limits, appropriate

warning messages are displayed during the operation. The results of the measurement

should be considered suspect until the exact nature of the problem is determined.

Examples of warning messages are described in the following paragraphs.

Reflected Power (Low or High)

Reflected power is the total amount of light (reflected plus scattered) reaching the photo-

detector. The main component is the light reflected from the probe/fluid interface, but it

includes the light scattered from the particles, which is normally no more than a few

percent of the reflected light level.

With clear water in the cell, the reflected power is nominally 3, but can vary considerably

due to minute variations in the probe/fluid interface caused by such things as a buildup of

thin layers of contamination on the probe face. Note: these values for reflected power

are inherent to the Nanotrac Wave system and software; there may be times during

operation that these values are not displayed to the user.

With fluids other than water, the expected reflected power is computed from the

refractive index of the fluid and is typically lower than the power for water.

Variations in reflected power do not affect the accuracy of the particle size measurement.

However, if the measured reflected power exceeds the limits for the fluid used, the

REFLECTED POWER LOW (or HIGH) message is displayed after a run, indicating the

need for corrective action. Possible causes of reflected power deviation and corrective

actions are these:

Dirty probe(s). Gently clean the cell and the optical probes.

No sample in the cell. Place sample in the Cell, or in the case of SetZero, place clean

fluid in the Cell, the same as used for sample mixtures.

Wrong refractive index value. Enter the correct value in the Flex 'Setup - Options -

Analysis - Fluid Information' screen.

Excessive particle concentration. Use properly diluted sample.

After cleaning the cell and probe or eliminating other potential causes, perform a

SETZERO and a run. If the REFLECTED POWER LOW (or HIGH) warning still

appears at the end of the run, other possible causes are:

Instrument malfunction

In this case, the Nanotrac Wave will require service or repair. Contact Microtrac

Technical Support for further information.

OM0022 Maintenance


Laser Power (Low or High)

Possible causes of large changes in laser power values are:

Laser degradation

Fault in laser drive circuitry

Unauthorized and potentially damaging adjustment of the LASER PWR setpoint.

In this case, the Nanotrac Wave will require service or repair. Contact Microtrac

Technical Support for further information.

Excessive Loading

During a RUN, reflected power level should not be more that 10% higher than the level

measured during SETZERO. Light scattered from the particles is normally no more than

a few percent of the total light arriving at the detector. As particle concentration

increases, scattered light increases and can become a significant portion of the total light.

Increased scattered light causes certain second-order effects, such as self-beating signals

and multiple scattering signals, to become apparent. Such second-order effects introduce

errors in the detected frequency distribution and subsequently in the reported particle

distribution. The typical effect is to give a distribution that is broader than normal and

skewed toward smaller particle sizes.

The onset of second-order effects is possible when a RUN reflected power level is 10%

above that reported during a SETZERO. In this case, the particle distribution is still

computed and presented, and the EXCESSIVE LOADING message is displayed during

the run. This is a warning that the reported distribution may contain errors and the

sample should be diluted and run again.

If this message then persists, contact Microtrac Technical Support for further

information.

Invalid Temperature

This message indicates that the sample cell temperature, as determined by the probe's

temperature sensor, is outside the +5oC to +90

oC design limits of the instrument. If the

user is neither cooling their sample below the low limit, nor heating their sample above

the high limit, then this error message is likely due to a hardware problem that the user

cannot fix. Additional diagnostic can be performed by running the Bench Status check,

previously described. Note that 'Cell Temperature' is indicated. If Bench Status is

performed with no liquid in the cell, then 'Cell Temperature' should give an approximate

indication of surrounding ambient temperature; for example a typical temperature-

controlled laboratory may show a readout of '22.0°C' or thereabouts. If this error persists,

and the Bench Status indicates abnormal ambient temperature, then contact Microtrac

Technical Support for repair or service information.

Unstable Temperature

The sample temperature must be known for an accurate computation of the particle size

distribution, since the viscosity of the dispersing fluid is a function of temperature. The

temperature is measured during a run. The average value is used to compute the fluid

viscosity. Temperature variations during the run add uncertainty to the viscosity and to

the computed particle distribution. The UNSTABLE TEMPERATURE warning message

OM0022 Maintenance


is displayed at the end of a run if the computed uncertainty exceeds +/-0.3 standard

(fourth-root) channel widths.

This error could be due to temperature fluctuations of the surrounding environments,

particularly during especially long run times. If possible, move the Nanotrac Wave to a

more temperature-stable area, or provide protection or shrouding to prevent excessive

temperature flow across the unit. Additionally, try running with the minimum

recommended run-time. If the error persists, contact Microtrac Technical Support for

further information.

Troubleshooting: Why Did The Temperature Control Fail to Start, or Why Did It Automatically Shut Down?

If the Wave analyzer is equipped with Temperature Control option, and if during a

Temperature Control operation, the Wave automatically shuts down the operation

without any user action, then something went wrong during the operation. Possible

reasons include:

* The Access Cover was opened; breaking the interlock. This will automatically cause

any Temperature operation to abort. In this case, close the Access Cover and the

Temperature operation can be started over.

* Some part of the Temperature Control system experienced a hardware failure, such as

overtemperature condition, broken temperature sensor, etc. In this case contact

Microtrac Technical Support for assistance.

Troubleshooting: Why Did The Temperature Not Reach The Setpoint?

If the temperature did not reach the desired setpoint, it could be due to several reasons:

* There is not enough airflow around the Wave unit to allow the Temperature Control

system to operate properly. Clear away obstructions that may be surrounding the

Wave, and try again.

* Temperature Control hardware has become worn or dirty with use over time. It may

be possible to recalibrate the hardware to offset this wear. Contact Microtrac for

assistance.

* Some part of the Temperature Control system experienced a hardware failure, such as

overtemperature condition, broken temperature sensor, etc. In this case contact

Microtrac Technical Support for assistance.

OM0022 Maintenance


5.5 Requesting Service

The Nanotrac Wave contains no user serviceable parts. Nanotrac Wave service or repair

should be coordinated by contacting Microtrac Technical Support:



Largo, FL 33773

(727) 507-9770

ATTENTION

The shipping cartons in which the Nanotrac Wave was shipped have been optimized for this instrument. Microtrac recommends that all cartons and shipping materials be retained, in the event that the Nanotrac Wave has to be returned to Microtrac for repair or service.

5.6 Requesting Parts and Accessories

Available accessories for the Nanotrac Wave may include (this list may be updated from

time to time, check with Microtrac for any changes):

Description

Standard reference material; sizing standard; polystyrene spheres

Standard reference material; zetapotential standard

Disposable pipette, fine tip

Microbrushes

Accessory kit (contains all of the above)

Chamois swabs

Parts and accessories can be ordered by contacting:

Microtrac Order Entry 148 Keystone Drive

Montgomeryville, PA 18936 TEL (215) 619-9920

FAX (215) 619-9932

www.microtrac.com

http://www.microtrac.com/